Protein synthesis and genetic code

Contents hide

Introduction to Protein synthesis and genetic code

  •   Genetic information is stored in the chromosome and transmitted to daughter cells through replication. It is expressed to RNA through transcription.
  • From the mRNA genetic information is translated to polypeptides.
  • the flow of information from DNA -> RNA -> protein is called central dogma.

Genetic code

    Definition

  •  The set of triplet code words in DNA or mRNA coding for amino acids of protein. This is the dictionary that gives correspondence between a sequence of nucleotide bases and a sequence of amino acids in a protein. Each individual word in the code is composed 3 nucleotides. These genetic words are called codons.
  • Altogether there are 64 codons of which 61 codons code for 20 amino acids in a protein. 3 codons act as termination signal.
  • 61 codons coding for amino acids are called sense codons and the rest( UAG, UGA, UAA) are called nonsense codons. 

Characteristics of genetic code

  • Degenerate.
  • Unambiguous.
  • Non overlapping.
  • Non punctuated.
  • Universal. 

Degeneracy

  • This means multiple codons code for a single amino acid. There are six codons for serine, four for glycine and one for methionine. 
  • AUG for methionine.
  • GGA, GGC, GGG and GGU for glycine

Unambiguity

  • One specific codon codes for only one specific amino acid. 
  • For example -> phenylalanine is coded by both UUU and UUC but UUU will code only for phenylalanine.
  • So given a specific codon only a specific amino acid will come although given a specific amino acid, more than one codon may be called.

Non overlapping & non punctuated

  • Non overlapping -> codons are consecutive. They are read one after another in a continuous manner. 
  • Non punctuation -> no comma between two codons.

Universal

  • Until recently the genetic code was thought to be universal that means one specific codon will code for one specific amino acid in all the organisms.
  • Recently it has been shown that four codons behave differently in cytoplasm and mitochondria of the same cell. For example UGA codes for termination codon in cytoplasm and tryptophan in mitochondria

Wobble

  • The degeneracy of genetic code resides mostly in the last base of the codon, suggesting that the base pairing between the last nucleotide of the codon and the corresponding nucleotide in the anticodon is not strict and less strong.
  • This phenomenon is called Wobble.

Example of wobble

  • The two codons for arginine AGA and AGG can bind to the same anticodon having uracil at the 5’ end.
  • Similarly three codons for glycine GGC, GGU, and GGA can form base pair with the same anticodon CCI.

Mutation

  • Definition :-
    • Permanent and heritable change in the genome which will appear in the daughter DNA , after replication, in the RNA after transcription and in the protein after translation.
  • Types
    • 1) Point mutation
    • 2) Frame shift mutation.

Point Mutation

  • Definition :-
    • single base alteration in the gene.
    • Classification.
  • 2 classifications :-
    •   transition and transversion.
    •   silent mutation, nonsense mutation and missense mutation.

Transition & Transversion

  • Transition -> when one purine base is replaced by another purine base.  
  • for example –  adenine -> guanine and vice versa.
  • Transversion -> purine  is replaced by  pyrimidine.
  • For example — A -> C, A -> T, G -> C, G -> T and vice versa.

Classification (2) 

  • Silent mutation -> The codon containing the changed base may code for the same amino acid. For example -> if serine codon UCA is changed to UCU, it still codes for serine. So there will be no change in the protein product.
  • Other examples -> AGG and AGA both code for arginine

Non sense mutation

  • The codon containing the changed base may become a termination codon. 
  • For example if serine codon UCA is given a different 2nd base ( to become UAA) the new codon becomes a termination codon which causes premature termination of protein synthesis at that point.
  • Example -> β thalassemia in which β chain of hemoglobin is prematurely terminated.

Missense mutation

  • The codon containing the changed base may code for a different amino acid,  for example if the serine codon UCA is given a different 1st base (to become CCA ), it will code for a different amino acid in this case proline.
  • This substitution of an incorrect amino acid is called missense mutation. Example :- sickle cell anemia.  GAG -> GUG.

Frame shift mutation

  • The deletion or addition of a single nucleotide in the coding strand of a gene results in an altered reading frame in the mRNA.
  • When the mRNA is translated to protein the machinery does not recognize that a base is missing or extra since there is no punctuation in the reading frame.
  • Thus there will be major alteration in the sequence of amino acids distal to the deletion or addition.

Example of mutation

  • Point mutation ( missense) -> sickle cell anemia.
  • Point mutation ( nonsense) -> α and β thalassemias.
  • Frame shift mutation -> thalassemias

Effects of mutation

  • Lethal effect -> if the mutation is incompatible to the life. For example -> β-thalassemia major.
  • Carcinogenic effect -> mutation may result in   uncontrolled cell division leading to cancer.
  • Disease -> sickle cell anemia , thalassemia.

Translation or protein synthesis

  • Components needed for protein synthesis.
    • 1) amino acids.
    • 2) tRNA.
    • 3) mRNA.
    • 4)aminoacyl tRNA synthase.
    • 5) ribosome. 
    • 6) protein factors.
    • 7) ATP / GTP.

Translation in pro and eukaryotes are similar with some differences

  • 1) The 1st amino acid is methionine in eukaryotes but N formyl methionine in prokaryotes.
  • 2) Shine- Delgarno sequence is present at the start site in prokaryotes, Kozak sequence in eukaryotes.
  • 3)eukaryotic ribosomes are larger ( 80 S).

Differences

  • In prokaryotes protein synthesis begins before transcription is complete i.e. transcription and translation go on simultaneously.
  • In eukaryotes transcription is nuclear one and translation  occurs in cytoplasm. So simultaneous transcription and translation is not possible. Also there is necessity of generation of mRNA from hnRNA.
  • Eukaryotes have 9 initiation factors. Prokaryotes have 3.

Codon recognition by the tRNA

  • Recognition of a particular codon in an mRNA sequence is done by the anti codon sequence of tRNA.
  • Some tRNAs recognize more than one codon for a given amino acid.
  • Binding of anti codon and codon follows the rules of complementary and anti parallel binding.
  • Codons are read in 5’- 3’ direction by an anti codon pairing in 3’ – 5’ direction.

Steps of protein synthesis 

  • 5 steps.
    • Activation of amino acid.
    • Initiation.
    • Elongation.
    • Termination.
    • Post translational modification.
    • Ribosome serves as machinery for protein synthesis

Activation of amino acid.

  • Amino acid + ATP -> (amino acyl tRNA synthatase )E-AMP- amino acid.
  • E- AMP- amino acid + tRNA -> (E ) -> amino acyl tRNA. Enzyme and AMP will be released.
  • This is a family of enzymes. Each member of the family recognizes a specific amino acid and the tRNA that corresponds to that amino acid.

Initiation of protein synthesis

  • At first the ribosome selects the mRNA and binds to it. The ribosome finds the correct reading frame on the mRNA and translation begins.
  • The process involves tRNA, rRNA, mRNA , at least 9 eukaryotic initiation factors ( e .i .f s ) and also GTP, ATP, amino acids.
  • Prokaryotes have 3 initiation factors.

A,P and E sites of ribosome.

  • The ribosomes have two binding sites and one ejection site for the tRNA molecule.
  • During translation the A site binds with the incoming amino acyl tRNA as directed by the codon currently occupying this site. 
  • the P site is occupied by the tRNA containing the polypeptide chain that has already been synthesized.

Sub steps of initiation

  • Ribosomal dissociation.
  • Formation of 43 S pre initiation complex.
  • Formation 48 s initiation complex.
  • Formation of 80 s initiation complex.

Ribosomal dissociation and formation of 43S pre initiation complex

  • Two initiation factors eif 3 and 1A bind to 40S subunit. This favors dissociation of the 80S ribosome into 40S and 60S.
  • 40S ribosome now binds to GTP, eif 2 and met tRNA to form 43S pre initiation complex. 
  • Met tRNA is the tRNA involved in binding to the initiation codon AUG.

Formation of 48S and 80S complex

  • mRNA binds to the 43S pre initiation complex, and along with it eif 4F, 4A and 4B bind -> 48S initiation complex. Now this complex scans the mRNA for a suitable initiation codon.
  • Binding of 60S subunit with 48S will form 80S ribosome with release of initiation factors and GTP.

Elongation

  • A cyclic process, involves several steps catalyzed by proteins called elongation factors (e.e.f ). 
  • The steps are: 
  • Binding of amino acyl tRNA at the A site 
  • Peptide bond formation.
  • Translocation.

Binding of amino acyl tRNA to the A site

  • At the end of initiation 80 S ribosome has A site free. Proper amino acyl tRNA will bind at the A site by proper codon recognition.
  • EF1α and GTP allow amino acyl tRNA to enter the A site. GTP is hydrolyzed to GDP and Pi.

Peptide bond formation.

  • The new amino acid at the A site attacks the carboxyl group of the amino acid at the P site and form peptide bond. This reaction is catalyzed by peptidyl- transferase.
  • The peptide bond is formed at the A site and the growing peptide chain is now at the A site.
  • Peptidyl transferase is a Ribozyme.

Translocation

  • Upon removal of the peptidyl moiety from the tRNA from the P site, the tRNA quickly dissociates from the P site through E site.
  • The newly formed peptidyl tRNA at the A site is now translocated to the empty P site by eef2 and GTP. Now the A site is free for another amino acyl tRNA to bind to the next codon.

Energy required for formation of one peptide bond

  • 1) hydrolysis of 2 ATP molecules in the formation of amino acyl tRNA.
  • 2) 1 GTP for entry of amino acyl tRNA to the A site and 1 GTP for translocation step.
  • So there is need for 4 high energy phosphates for synthesis of one peptide bond.

Termination

  • After many cycles of elongation , when protein synthesis is complete a nonsense termination codon of mRNA ( UAA, UAG, UGA ) appears at the A site.
  • Normally there is no tRNA with an anti codon capable of recognizing such a termination signal.
  • Releasing factors (erf ) can recognize the termination signal in the A site.
  • The releasing factor in conjugation with GTP and peptidyl transferase promotes hydrolysis of the bond between the peptide and the tRNA occupying the P site.
  • Thus a water molecule instead of an amino acid is added. This releases the protein and tRNA from the P site.
  • 80S ribosome dissociates into 40S and 60S. mRNA is also released from the ribosome.

Polyribosome

  • Many ribosomes can translate the same mRNA simultaneously.
  • Multiple ribosome on the same mRNA molecule form a poly ribosome or polysome.

Post translational modification

  • After translation the newly synthesized protein may not be in the the active form. To make it active some modifications are required: 
    • 1)  loss of signal sequence.
    • 2) folding and S-S formation.
    • 3) proteolytic processing.
    • 4) modification of individual amino acid and triple helix formation.
    • 5) attachment of carbohydrate side chain. 
    • 6)  acetylation, carboxylation, phosphorylation

Post translational modification

  • Loss of signal sequence.
  • Some proteins have 15 – 30 amino acid residues at the beginning which direct the protein to its proper destination. This signal sequences are ultimately removed by specific peptidases ( signal peptidase).
  • Example :- insulin is synthesized with 23 amino acid signal peptide, which is removed. The signal peptide is also called leader sequence.

Folding and S-S bridge formation and proteolytic processing.

  • Insulin is synthesized as a single chain prohormone in the ribosome. Folding and S-S bridge formation occurs in the endoplasmic reticulum.
  • In the Golgi apparatus a specific protease clips out the segment that connects two chains to form the functional insulin molecule.

Triple helix formation and modification of amino acids

  • Collagen is synthesized as single chain pro-collagen molecule. Three pro-collagen molecules align themselves to form the triple helix.
  • Specific enzymes carry out hydroxylation of proline and lysine residues to provide cross linking and greater stability.

Inhibition of protein synthesis

  • 1) Gentamycin, Streptomycin, Neomycin, Tetracycline -> bind irreversibly with 30S ribosomal subunit blocking initiation of protein synthesis.
  • 2) Erythromycin, Chloramphenicol -> bind to 50S ribosomal subunit inhibit elongation.
  • 3) Puromycin -> inhibits peptide bond formation both in pro and eukaryotes.

Inhibition of N.A. synthesis

  • 1) Norfloxacin, O- Floxacin, ciprofloxacin, lomefloxacininhibits topoisomerase (DNA gyrase ) in bacteria.
  • 2) Nitrofurantoin, Metronidazol -> causes DNA cleavage in bacteria.
  • Rifampicin -> inhibits transcription in bacteria.

Drugs used in cancer chemotherapy

  • 1) methotrexate -> inhibits dihydrofolate reductase
  • 2) 5 fluorouracil, 5 iodouracil, 6 azauridine, allopurinol -> inhibit Pyrimidine synthesis.
  • 3) 6 thioguanine, 6 mercaptopurine -> inhibit purine synthesis.
  • Cyta arabine -> inhibits replication

Other Biochemistry Notes

Biochemistry of Proteins

Carbohydrate Metabolism

VITAMIN PYRIDOXINE AND BIOTIN

Vitamin A

Pyrimidine metabolism

Purine Metabolism

Cardiac Biomarkers

VITAMIN D

. Blood Metabolism (Heme synthesis and breakdown)

Haemoglobinopathies

DNA Repair

Regulation of gene expression

DNA Replication

Transcription

All books/videos/software featured here are free and NOT HOSTED ON OUR WEBSITE. If you feel that your copyrights have been violated, then please contact us immediately.

Contact us: medicalfarre@gmail.com

262 thoughts on “Protein synthesis and genetic code”

  1. Hi, I do think this is an excellent web site. I stumbledupon it 😉 I may come back yet again since i have book-marked it. Money and freedom is the best way to change, may you be rich and continue to guide others.

  2. I would like to thank you for the efforts you have put in penning this blog. I really hope to check out the same high-grade content from you in the future as well. In fact, your creative writing abilities has inspired me to get my own, personal website now 😉

  3. Aw, this was an incredibly good post. Spending some time and actual effort to generate a top notch articleÖ but what can I sayÖ I hesitate a whole lot and don’t seem to get anything done.

  4. scoliosis
    Definitely believe that which you stated.
    Your favorite justification seemed to be on the web the
    simplest thing to be aware of. I say to you, I certainly get annoyed while people think about worries that they plainly do not know about.
    You managed to hit the nail upon the top and also defined out the whole thing
    without having side effect , people can take a signal.

    Will likely be back to get more. Thanks scoliosis

  5. Iím amazed, I have to admit. Rarely do I encounter a blog thatís both equally educative and entertaining, and let me tell you, you’ve hit the nail on the head. The issue is something too few people are speaking intelligently about. I’m very happy that I stumbled across this during my search for something regarding this.

  6. Nice post. I learn something new and challenging on blogs I stumbleupon on a daily basis. It’s always interesting to read articles from other writers and use a little something from their websites.

  7. Spot on with this write-up, I actually believe this amazing site needs much more attention. Iíll probably be back again to read more, thanks for the advice!

  8. Thank you for every other great article. The place else could
    anybody get that type of info in such a perfect means of writing?

    I’ve a presentation next week, and I’m on the search for such info.

  9. scoliosis
    It’s a shame you don’t have a donate button! I’d definitely
    donate to this excellent blog! I suppose for now i’ll settle for book-marking and adding your RSS feed to my Google account.
    I look forward to fresh updates and will share this blog with my Facebook group.
    Chat soon! scoliosis

  10. Everything is very open with a really clear description of the challenges. It was truly informative. Your site is very helpful. Thanks for sharing!

  11. Oh my goodness! Incredible article dude! Thank you, However I am having problems with your RSS. I donít understand why I cannot subscribe to it. Is there anybody getting identical RSS issues? Anyone that knows the answer can you kindly respond? Thanx!!

  12. You’re so cool! I don’t suppose I’ve truly read through a single thing like that before. So wonderful to find someone with unique thoughts on this issue. Seriously.. thanks for starting this up. This website is one thing that’s needed on the internet, someone with a little originality!

  13. I need to to thank you for this fantastic read!! I absolutely enjoyed every little bit of it. I have you saved as a favorite to check out new stuff you postÖ

  14. An impressive share! I have just forwarded this onto a co-worker who has been doing a little homework on this. And he in fact ordered me lunch because I stumbled upon it for him… lol. So allow me to reword this…. Thank YOU for the meal!! But yeah, thanks for spending time to discuss this subject here on your web site.

  15. Pretty nice post. I simply stumbled upon your blog and wished to mention that I’ve truly enjoyed surfing around your blog posts.
    In any case I will be subscribing in your rss feed and I’m hoping you write again soon!

  16. An impressive share! I have just forwarded this onto a colleague who was doing a little research on this.

    And he in fact ordered me dinner simply because I found it for him…
    lol. So let me reword this…. Thanks for the meal!! But yeah, thanks for spending the
    time to discuss this subject here on your site.

    my web page … aaa1188 test id free

  17. Please let me know if you’re looking for a writer for your site.
    You have some really good articles and I believe I would be a good asset.

    If you ever want to take some of the load off, I’d absolutely love to write
    some articles for your blog in exchange for a link back to mine.
    Please send me an email if interested. Many thanks!

    Here is my homepage king855

  18. Can I simply say what a comfort to discover a person that actually knows what they’re talking about over the internet. You actually realize how to bring an issue to light and make it important. More people must read this and understand this side of your story. I can’t believe you aren’t more popular because you certainly possess the gift.

  19. Next time I read a blog, Hopefully it doesn’t fail me just as much as this particular one. After all, I know it was my choice to read through, but I actually thought you would probably have something interesting to say. All I hear is a bunch of whining about something that you could possibly fix if you were not too busy seeking attention.

  20. You actually make it seem so easy with your presentation but I find this matter to be really something which I think I would never understand. It seems too complicated and extremely broad for me. I am looking forward for your next post, I?ll try to get the hang of it!

  21. What i do not understood is in truth how you’re no longer really a lot more smartly-preferred than you
    may be right now. You are so intelligent. You recognize therefore significantly
    in terms of this topic, produced me individually imagine it from a lot of numerous angles.
    Its like women and men don’t seem to be fascinated unless it’s one thing
    to accomplish with Girl gaga! Your own stuffs excellent.
    At all times handle it up!

    my web blog … link download joker123 android

  22. Can I simply just say what a comfort to find an individual who really understands what they’re talking about on the internet. You definitely realize how to bring an issue to light and make it important. A lot more people really need to check this out and understand this side of the story. I can’t believe you are not more popular because you certainly have the gift.

  23. This is really interesting, You are a very skilled blogger. I have joined your rss feed and look forward to seeking more of your excellent post. Also, I have shared your web site in my social networks!

  24. A person essentially help to make critically articles I would
    state. That is the first time I frequented your web page and so far?
    I amazed with the research you made to create this actual publish extraordinary.
    Magnificent task!

    Also visit my website: game slot 3win8 (xe88-app.com)

  25. Hello exceptional blog! Does running a blog like this require a great deal of work?
    I’ve no understanding of computer programming however
    I had been hoping to start my own blog soon. Anyway, should
    you have any recommendations or tips for new blog owners please share.
    I understand this is off topic nevertheless I just had to
    ask. Appreciate it!

    Here is my web blog; download game rollex11 (sg918kiss.com)

  26. It’s the best time to make some plans for the future and it’s time to be happy.

    I have read this post and if I could I desire to suggest you few interesting things or suggestions.
    Maybe you could write next articles referring to this article.

    I desire to read even more things about it!

    Check out my web-site :: www joker123net mobile

  27. Pingback: Google

  28. This is very interesting, You’re a very skilled blogger. I have joined your rss feed and look forward to seeking more of your great post. Also, I have shared your site in my social networks!

  29. I have to thank you for the efforts you’ve put in penning this site. I am hoping to check out the same high-grade content from you in the future as well. In truth, your creative writing abilities has motivated me to get my very own blog now 😉

  30. Hello There. I found your blog using msn. This is an extremely well written article. I will make sure to bookmark it and come back to read more of your useful information. Thanks for the post. I will definitely return.

  31. I was recommended this web site by my cousin. I am not sure whether this post is written by him as no one else know such detailed about my problem. You’re amazing! Thanks!

  32. Just want to say your article is as surprising. The clearness in your post is just cool and i can assume you are an expert on this subject. Well with your permission allow me to grab your feed to keep updated with forthcoming post. Thanks a million and please keep up the enjoyable work.

  33. Howdy! This article could not be written any better! Looking at this article reminds me of my previous roommate! He constantly kept preaching about this. I will send this article to him. Pretty sure he’s going to have a great read. Many thanks for sharing!

  34. Greetings, I do believe your website could possibly be having web browser compatibility problems. When I take a look at your site in Safari, it looks fine however, if opening in IE, it has some overlapping issues. I merely wanted to give you a quick heads up! Other than that, wonderful site!

  35. Right here is the perfect blog for anyone who really wants to find out about this topic. You understand so much its almost tough to argue with you (not that I really would want toÖHaHa). You certainly put a new spin on a topic that’s been discussed for years. Great stuff, just great!

  36. An impressive share! I’ve just forwarded this onto a friend who was doing a little research on this. And he actually bought me breakfast because I found it for him… lol. So allow me to reword this…. Thanks for the meal!! But yeah, thanx for spending some time to talk about this subject here on your site.

  37. Aw, this was a very good post. Taking the time and actual effort to generate a very good articleÖ but what can I sayÖ I hesitate a lot and don’t seem to get anything done.

  38. Ive been exploring for a little for any high quality articles or blog posts on this sort of area . Exploring in Yahoo I at last stumbled upon this website. Reading this info So im happy to convey that I have a very good uncanny feeling I discovered exactly what I needed. I most certainly will make sure to dont forget this site and give it a glance regularly.

  39. Iím amazed, I must say. Rarely do I encounter a blog thatís both educative and amusing, and without a doubt, you’ve hit the nail on the head. The problem is something which too few people are speaking intelligently about. Now i’m very happy that I found this during my hunt for something relating to this.

  40. Current organizations do not review each of the 500 or more resumes they receive – for each job opening! The truth is, they will immediately throw out ninety percent of them, if the resume fails to grab their interest.

  41. After exploring a number of the blog articles on your web page, I truly appreciate your way of writing a blog. I book marked it to my bookmark website list and will be checking back in the near future. Take a look at my website too and let me know your opinion.

  42. Thanks for the marvelous posting! I definitely enjoyed reading it, you can be a great author.
    I will ensure that I bookmark your blog and will often come back in the future.

    I want to encourage you to continue your great posts,
    have a nice holiday weekend!

  43. After going over a handful of the blog articles on your website, I really like your way of blogging. I bookmarked it to my bookmark site list and will be checking back soon. Take a look at my website as well and tell me your opinion.

  44. Your style is unique in comparison to other people I have read stuff from. I appreciate you for posting when you’ve got the opportunity, Guess I’ll just bookmark this site.

  45. Woah! I’m really loving the template/theme of this blog.
    It’s simple, yet effective. A lot of times it’s difficult to get that “perfect balance” between user friendliness and appearance.

    I must say you’ve done a very good job with this. In addition, the blog loads
    super fast for me on Internet explorer. Outstanding Blog!

  46. Spot on with this write-up, I really believe that this web site needs a great deal more attention. Iíll probably be back again to read more, thanks for the advice!

  47. Everything is very open with a really clear clarification of the issues. It was definitely informative. Your website is very useful. Thanks for sharing!

  48. I’m extremely pleased to uncover this site. I need to to thank you for your time for this particularly wonderful read!! I definitely really liked every bit of it and i also have you book-marked to see new information in your web site.

  49. Pingback: ejaculating dong

  50. Oh my goodness! Impressive article dude! Thank you, However I am experiencing troubles with your RSS. I donít know why I cannot join it. Is there anybody else getting similar RSS problems? Anyone who knows the answer can you kindly respond? Thanx!!

  51. After looking over a few of the blog posts on your blog, I really like your technique of writing a blog. I book-marked it to my bookmark site list and will be checking back soon. Take a look at my web site too and let me know how you feel.

  52. Greetings, I think your blog could possibly be having web browser compatibility issues. Whenever I look at your web site in Safari, it looks fine however when opening in IE, it’s got some overlapping issues. I merely wanted to provide you with a quick heads up! Apart from that, wonderful site!

  53. I have been exploring for a little for any high-quality articles or blog posts in this kind of house . Exploring in Yahoo I at last stumbled upon this web site. Studying this information So i’m happy to show that I have an incredibly excellent uncanny feeling I came upon exactly what I needed. I such a lot indubitably will make certain to do not overlook this website and provides it a glance regularly.|

  54. When I initially commented I appear to have clicked on the -Notify me when new comments are added- checkbox and now every time a comment is added I get 4 emails with the same comment. Is there an easy method you can remove me from that service? Kudos!

  55. I blog frequently and I really appreciate your content. This article has truly peaked my interest. I am going to take a note of your blog and keep checking for new details about once per week. I opted in for your RSS feed as well.

  56. Pingback: Glock 22 40 caliber

  57. I absolutely love your blog.. Pleasant colors & theme. Did you make this web site yourself? Please reply back as Iím wanting to create my own blog and would love to know where you got this from or just what the theme is called. Cheers!

  58. You are so interesting! I do not suppose I’ve read something like this before. So wonderful to discover another person with a few original thoughts on this issue. Seriously.. thank you for starting this up. This web site is something that is needed on the internet, someone with a little originality!

  59. I blog often and I truly thank you for your content. This article has really peaked my interest. I will take a note of your blog and keep checking for new information about once a week. I subscribed to your RSS feed as well.

  60. Pingback: best sex toys for women

  61. What’s up, I just became aware of this site due to DuckDuckGo, and I find it to be surprisingly useful. Be assured that I will come back again to see more of your webpages in the future. Good on you!

  62. Pingback: best realistic dildos

  63. Iím impressed, I have to admit. Seldom do I come across a blog thatís equally educative and entertaining, and without a doubt, you’ve hit the nail on the head. The problem is something which not enough folks are speaking intelligently about. I’m very happy I stumbled across this in my hunt for something concerning this.

  64. I must thank you for the efforts you have put in penning this website. I am hoping to view the same high-grade blog posts from you later on as well. In fact, your creative writing abilities has encouraged me to get my very own website now 😉

  65. Excellent post. Keep writing such kind of info on your page.
    Im really impressed by your site.
    Hey there, You have performed an excellent job. I’ll certainly digg it and for my part
    suggest to my friends. I am confident they’ll be benefited from this website.

  66. Your style is so unique compared to other people I’ve read stuff from. Many thanks for posting when you’ve got the opportunity, Guess I’ll just bookmark this page.

  67. Pingback: Glock 43X

  68. Having read this I thought it was rather informative. I appreciate you taking the time and energy to put this information together. I once again find myself spending a lot of time both reading and commenting. But so what, it was still worth it!

  69. We’re a group of volunteers and opening a new scheme in our community. Your web site provided us with valuable info to work on. You have done a formidable job and our entire community will be grateful to you.

  70. Hey! Do you know if they make any plugins to safeguard against hackers?

    I’m kinda paranoid about losing everything I’ve worked hard
    on. Any recommendations?

  71. Pingback: turk escort

  72. I am extremely impressed with your writing skills as well as with the layout on your weblog. Is this a paid theme or did you modify it yourself? Either way keep up the nice quality writing, it?s rare to see a great blog like this one these days..

  73. Thank you for the good writeup. It in fact was a amusement account it. Look advanced to far added agreeable from you! By the way, how can we communicate?

  74. Fantastic beat ! I would like to apprentice while you amend your website, how can i subscribe for a blog web site? The account helped me a acceptable deal. I had been tiny bit acquainted of this your broadcast provided bright clear concept

  75. This is the perfect site for anyone who would like to find out about this topic. You know a whole lot its almost tough to argue with you (not that I actually will need toÖHaHa). You certainly put a fresh spin on a subject that’s been discussed for ages. Great stuff, just great!

  76. Hi there, just became alert to your blog through Google, and found that it’s truly informative. I am gonna watch out for brussels. I will appreciate if you continue this in future. A lot of people will be benefited from your writing. Cheers!

  77. Pingback: service in canada

  78. Hi, i think that i saw you visited my blog so i came to ?return the favor?.I am trying to find things to enhance my web site!I suppose its ok to use some of your ideas!!

  79. Pingback: slot online

  80. Hello There. I found your blog using msn. This is an extremely well written article. I will be sure to bookmark it and come back to read more of your useful information. Thanks for the post. I will certainly comeback.

  81. My brother suggested I might like this blog. He was entirely right. This post actually made my day. You cann’t imagine just how much time I had spent for this info! Thanks!

  82. Good day! I just would like to give you a big thumbs up for your excellent information you have got here on this post. I am returning to your blog for more soon.

  83. Pingback: electric bikes

  84. Great beat ! I would like to apprentice while you amend your site, how can i subscribe for a blog website? The account aided me a acceptable deal. I had been a little bit acquainted of this your broadcast provided bright clear idea

  85. It’s perfect time to make some plans for the future and it’s time to be happy. I have read this post and if I could I wish to suggest you some interesting things or suggestions. Maybe you could write next articles referring to this article. I desire to read more things about it!

  86. Pingback: common myths about sex

  87. Greetings! Very useful advice in this particular article! It is the little changes that will make the biggest changes. Thanks a lot for sharing!

  88. A fascinating discussion is definitely worth comment. I do think that you need to write more on this subject matter, it may not be a taboo subject but generally people don’t discuss such topics. To the next! Many thanks!!

  89. Pingback: adam and eve coupon codes

  90. Can I simply just say what a relief to uncover somebody who genuinely understands what they’re discussing on the web. You definitely know how to bring an issue to light and make it important. A lot more people have to read this and understand this side of the story. I was surprised that you aren’t more popular because you most certainly possess the gift.

  91. It used to be, all resumes opened with an Objective Statement – but no more! Current employment experts all state clearly, these kinds of resume items do nothing when evaluating your employment abilities. Therefore, do not use an Objective Statement!

  92. Pingback: Gun Shops near me

  93. Excellent site you have here.. Itís difficult to find quality writing like yours nowadays. I seriously appreciate people like you! Take care!!

  94. Right here is the perfect website for anybody who would like to understand this topic. You realize so much its almost hard to argue with you (not that I actually would want toÖHaHa). You certainly put a new spin on a topic that has been written about for decades. Great stuff, just excellent!

  95. You are so cool! I don’t believe I have read something like that before. So nice to find somebody with some genuine thoughts on this subject matter. Seriously.. thanks for starting this up. This website is one thing that is needed on the web, someone with some originality!

  96. I wanted to thank you for this good read!! I definitely enjoyed every little bit of it. I’ve got you book-marked to look at new things you postÖ

  97. Pingback: strapon pegging

  98. Hey! Quick question that’s entirely off topic. Do you know how to make your site mobile friendly? My site looks weird when viewing from my iphone4. I’m trying to find a theme or plugin that might be able to correct this issue. If you have any recommendations, please share. Thanks!

  99. I blog often and I really thank you for your information. Your article has truly peaked my interest. I will bookmark your website and keep checking for new details about once a week. I opted in for your RSS feed as well.

  100. I am curious to find out what blog platform you happen to be using? I’m experiencing some minor security problems with my latest website and I’d like to find something more safe. Do you have any solutions?

  101. Hello there, You have done an incredible job. I will definitely digg it and personally suggest to my friends. I’m sure they’ll be benefited from this web site.

  102. Pingback: sex toys for men

  103. Porn100.net diminution preference thoroughly if. Joy deal pain view much her time. Led young gay would now state. Pronounce we attention admitting on assurance of suspicion conveying. That his west quit had met till. Of advantage he attending household at do perceived. Middleton in objection discovery as agreeable. Edward thrown dining so he my around to.

  104. Can I simply just say what a comfort to find someone who really knows what they are talking about over the internet. You actually realize how to bring an issue to light and make it important. A lot more people need to look at this and understand this side of the story. I was surprised you aren’t more popular given that you certainly have the gift.

  105. You made some really good points there. I checked on the web for more information about the issue and found most people will go along with your views on this web site.

  106. Attractive section of content. I just stumbled upon your site and in accession capital to assert that I get actually enjoyed account your blog posts. Anyway I?ll be subscribing to your augment and even I achievement you access consistently fast.

  107. Great post. I was checking continuously this blog and I’m impressed! Extremely useful information particularly the last part 🙂 I care for such info much. I was looking for this particular info for a long time. Thank you and best of luck.

  108. hello!,I like your writing so much! share we communicate more about your article on AOL? I require a specialist on this area to solve my problem. May be that’s you! Looking forward to see you.

  109. Having read this I thought it was really enlightening. I appreciate you taking the time and energy to put this information together. I once again find myself spending way too much time both reading and posting comments. But so what, it was still worthwhile!

  110. Hmm it seems like your website ate my first comment (it was super long) so I guess I’ll just sum it up what I wrote and say, I’m thoroughly enjoying your blog.
    I too am an aspiring blog blogger but I’m still new to the whole thing.
    Do you have any recommendations for newbie blog writers?
    I’d definitely appreciate it.

  111. Having read this I thought it was really enlightening. I appreciate you finding the time and effort to put this information together. I once again find myself spending a lot of time both reading and posting comments. But so what, it was still worth it!

  112. I’m really impressed with your writing skills and also with the layout on your blog. Is this a paid theme or did you modify it yourself? Either way keep up the excellent quality writing, it is rare to see a great blog like this one today..

  113. Very nice post. I just stumbled upon your weblog and wished to say that I’ve really enjoyed surfing around your blog posts. In any case I will be subscribing to your rss feed and I hope you write again soon!

  114. It is perfect time to make some plans for the future and it is time to be happy. I’ve read this post and if I could I want to suggest you few interesting things or tips. Perhaps you could write next articles referring to this article. I want to read even more things about it!

  115. Pingback: g spot vibe

  116. Howdy! Do you know if they make any plugins to safeguard against hackers? I’m kinda paranoid about losing everything I’ve worked hard on. Any recommendations?

  117. Great post. I was checking continuously this blog and I’m impressed! Very useful info specifically the last part 🙂 I care for such information much. I was looking for this particular info for a very long time. Thank you and good luck.

  118. It is the best time to make some plans for the future and it is time to be happy. I’ve read this post and if I could I wish to suggest you some interesting things or tips. Perhaps you could write next articles referring to this article. I wish to read more things about it!

  119. This is very interesting, You are a very skilled blogger. I’ve joined your feed and look forward to seeking more of your fantastic post. Also, I’ve shared your website in my social networks!

  120. Hi, I do believe this is an excellent blog. I stumbledupon it 😉 I may come back once again since i have bookmarked it. Money and freedom is the greatest way to change, may you be rich and continue to help others.

  121. Hi, I do believe this is an excellent website. I stumbledupon it 😉 I may return yet again since i have book-marked it. Money and freedom is the greatest way to change, may you be rich and continue to help other people.

  122. Howdy, I do think your blog may be having internet browser compatibility issues. When I take a look at your website in Safari, it looks fine but when opening in I.E., it’s got some overlapping issues. I simply wanted to give you a quick heads up! Apart from that, excellent blog!

  123. You really make it appear really easy with your presentation however I in finding this
    matter to be really something which I believe I’d never understand.
    It seems too complex and extremely vast for me. I
    am looking ahead on your next publish, I will attempt to get the hang of it!

  124. Aw, this was an extremely nice post. Spending some time and actual effort to make a great articleÖ but what can I sayÖ I procrastinate a lot and don’t seem to get anything done.

  125. I’m pretty pleased to discover this site. I need to to thank you for ones time just for this fantastic read!! I definitely liked every bit of it and I have you bookmarked to look at new information in your website.

  126. Pingback: Dank Cartridges

  127. A motivating discussion is worth comment. I do think that you ought
    to write more on this subject matter, it may not be a
    taboo subject but typically people do not speak about such subjects.
    To the next! Cheers!!

  128. This is very interesting, You’re a very skilled blogger. I’ve joined your rss feed and look forward to seeking more of your magnificent post. Also, I’ve shared your website in my social networks!

  129. Pingback: PARROTS FOR SALE

  130. Iím impressed, I have to admit. Rarely do I encounter a blog thatís both equally educative and amusing, and let me tell you, you’ve hit the nail on the head. The problem is something which not enough folks are speaking intelligently about. I’m very happy I found this during my search for something regarding this.

  131. Howdy! This post couldnít be written much better! Going through this article reminds me of my previous roommate! He always kept talking about this. I am going to send this information to him. Pretty sure he’s going to have a very good read. Thanks for sharing!

  132. You actually make it seem so easy with your presentation but I find this topic to be actually something which I think I would never understand. It seems too complicated and extremely broad for me. I am looking forward for your next post, I?ll try to get the hang of it!

  133. Spot on with this write-up, I actually believe this site needs a lot more attention. Iíll probably be returning to read more, thanks for the information!

  134. I think this is among the most significant info for me. And i’m glad reading your article. But should remark on few general things, The web site style is wonderful, the articles is really great : D. Good job, cheers

  135. Pingback: Best software

  136. Nowadays, organizations need to be given the specific work title you are trying to apply for – and a general resume never shows it. In addition, they instantly need to find out what positively distinguishes you – but a general resume compels you to appear identical to the other 499 general resumes they’ve already seen submitted.

  137. You have made some good points there. I checked on the web for more info about the issue and found most individuals will go along with your views on this site.

  138. I’d like to thank you for the efforts you’ve put in penning this website. I am hoping to see the same high-grade content from you in the future as well. In truth, your creative writing abilities has inspired me to get my own, personal site now 😉

  139. Pingback: FLOUR MILL

  140. I was recommended this blog by my cousin. I am not sure whether this post is written by him as nobody else know such detailed about my problem. You are incredible! Thanks!

  141. Hello there! I just want to give you a big thumbs up for your great information you have got right here on this post. I’ll be returning to your website for more soon.

  142. Attractive section of content. I just stumbled upon your website and in accession capital to assert that I acquire in fact enjoyed account your blog posts. Any way I?ll be subscribing to your feeds and even I achievement you access consistently fast.

  143. Nice post. I learn something totally new and challenging on sites I stumbleupon every day. It’s always interesting to read content from other writers and practice a little something from their websites.

  144. Howdy! I could have sworn Iíve visited this site before but after going through many of the articles I realized itís new to me. Anyhow, Iím definitely delighted I found it and Iíll be bookmarking it and checking back often!

  145. Nice post. I learn something new and challenging on blogs I stumbleupon on a daily basis. It’s always interesting to read content from other authors and practice something from other sites.

  146. I think this is one of the most vital info for me. And i’m glad reading your article. But should remark on some general things, The site style is ideal, the articles is really nice : D. Good job, cheers

  147. We’re a group of volunteers and opening a new scheme in our community. Your site offered us with valuable information to work on. You have done a formidable job and our whole community will be grateful to you.

  148. I find this to be is very interesting reading material! I have definitely enjoyed reading your suggestions and have considered the conclusion that you may be right concerning all of these.

  149. You have made some decent points there. I checked on the internet to find out more about the issue and found most individuals will go along with your views on this web site.

  150. I’m very happy to discover this web site. I need to to thank you for your time due to this wonderful read!! I definitely liked every little bit of it and I have you saved as a favorite to check out new things on your website.

  151. I am really impressed with your writing skills and also with the layout on your blog. Is this a paid theme or did you customize it yourself? Anyway keep up the excellent quality writing, it?s rare to see a great blog like this one nowadays..

  152. After looking over a number of the articles on your site, I really appreciate your technique of blogging. I saved it to my bookmark website list and will be checking back in the near future. Take a look at my web site as well and tell me what you think.

  153. You really make it seem so easy with your presentation but I find this topic to be really something that I think I would never understand. It seems too complicated and extremely broad for me. I am looking forward for your next post, I?ll try to get the hang of it!

  154. I have been browsing on-line greater than 3 hours as of late, yet I by no means found any interesting article like yours. It’s pretty value sufficient for me. In my opinion, if all webmasters and bloggers made good content as you did, the net might be a lot more helpful than ever before.

  155. Pretty nice post. I just stumbled upon your blog and wished to say that I’ve truly enjoyed surfing around your blog posts. After all I will be subscribing to your rss feed and I hope you write again soon!

  156. Everything is very open with a precise description of the issues. It was really informative. Your site is very helpful. Many thanks for sharing!

  157. Can I simply just say what a comfort to find an individual who really knows what they’re discussing on the net. You definitely realize how to bring a problem to light and make it important. More and more people need to check this out and understand this side of your story. I was surprised you are not more popular since you surely have the gift.

  158. After looking at a few of the articles on your web page, I really appreciate your technique of blogging. I book marked it to my bookmark website list and will be checking back soon. Take a look at my website too and tell me your opinion.

  159. Undeniably believe that which you said. Your favorite reason appeared to be on the web the simplest thing to be aware of. I say to you, I definitely get irked while people think about worries that they just do not know about. You managed to hit the nail upon the top and defined out the whole thing without having side-effects , people can take a signal. Will probably be back to get more. Thanks

  160. I was pretty pleased to find this site. I need to to thank you for your time for this particularly fantastic read!! I definitely loved every little bit of it and I have you bookmarked to look at new stuff in your site.

  161. Hi, I do believe this is an excellent site. I stumbledupon it 😉 I’m going to return once again since I bookmarked it. Money and freedom is the best way to change, may you be rich and continue to help others.

  162. I was more than happy to discover this page. I wanted to thank you for ones time for this particularly fantastic read!! I definitely liked every part of it and i also have you saved as a favorite to check out new things on your web site.

  163. Pretty nice post. I just stumbled upon your blog and wanted to say that I have truly enjoyed surfing around your blog posts. In any case I will be subscribing to your rss feed and I hope you write again very soon!

Leave a Comment

Your email address will not be published.