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Post-Translational Modifications (PTMs)...

glycosylation

methylation

Protein diversity is essential in carrying out many cellular functions.

cell to cell signaling

enzymes

cell structure

The diversity of proteins greatly exceeds the number of proteins predicted by DNA coding. mRNA splicing at the transcriptional level and post-translational modifications further diversify the proteome.

Post-Translational Covalent

Modification of Proteins

Glycosylation

Glycoproteins

  • glycosylation mainly occurs in ER and Golgi apparatus
  • the mass of the carbohydrate can account for 1% - 80% of glycoprotein mass
  • common glycosylation involves covalent attachment of a complex oligosaccharide at Asparagine residue

Protein Glycosylation at Asparagine Residue

amide group of Asn residue acts as nucleophile to form N-glycosidic linkage

Summary

  • Post-translational modifications lead to the diversity in the proteome.
  • Post-translational modification is a chemical modification of a protein after its translation.
  • The different groups (i.e. methyls, carbohydrates, phosphoryls) that are get attached to protein lead to that protein's function within the cell.
  • Modifications can occur at amino acid side chains or at the polypeptide backbone.
  • Modifications (phosphorylations/dephosphorylations) of enzymes greatly influence its behavior/activity.

Cleavage at Peptide Bonds (Proteolytic Cleavage)

  • Insulin is synthesized by peptide cleavages
  • Most proteins undergo proteolytic cleavage following translation (simplest form is removal of initiatior AA Met and/or removal of signal sequence)

1. Pre-proinsulin is synthesized from insulin gene and directed into the ER by the signal sequence where it folds into proper conformation and is stabilized by 3 disulfide bonds.

  • activation of proenzymes occur in digestive enzymes, blood clotting cascade

2. The signal sequence is removed, and proinsulin is further processed in the Golgi apparatus, where the C-peptide is removed

occur at amino acid side chains or peptide linkages

are performed by 5% of the proteins in proteome

can occur at any step of the "life cycle" of the protein

Enzyme dependent covalent addition of a chemical group, usually an electrophilic fragment of cosubstrate to a side chain residue of protein. Side chain modified is usually electron rich, acting as the nucleophile.

Covalent cleavage of peptide backbone by proteases or less commonly, by autolytic cleavage.

transferases

phosphatases

right after translation

later on

phosphorylation

kinases

acetylation

Methylation

SAM

Phosphorylation

  • occurs on nitrogen (irreversible) or oxygen atoms (reversible)
  • reversible protein phosphorylation occurs on

serine, threonine or tyrosine residues

Phosphorylation Mechanism

  • the phosphoryl transfer is facilitated by Mg

2+

  • most common methylation occurs at the E-amine of lysine and arginine residues
  • phosphorylation regulates protein function (activation or deactivation) & cell signaling by causing conformational changes
  • When hepatocytes have high levels of glucagon, pyruvate kinase is phosphorylated by protein kinase A. This phosphorylation decreases activity of pyruvate kinase.

Methylation by S-adenosyl methionine (SAM)

  • methylation occurs with the help of methyltransferases
  • residues can be progressively methylated
  • Histones (proteins that package and order DNA in structural units) can act epigenetically to repress or activate gene expression. In H3, residues can be mono, di, and tri-methylated.
  • methylation is mediated by methyltransferases and S-adenosyl methionine (SAM) is primary methyl group donor
  • these PTM's play critical roles in regulation of cellular processes (i.e. activation/deactivation of enzymes)
  • methylation is well-known mechanism of epigenetic regulation (histone methylation and demethylation influences the availability of DNA for transcription)
  • kinases and phosphorylases make up the largest class of PTM enzymes
  • methylation occurs so often that SAM has been suggested to be the most used substrate in enzymatic reactions after ATP!

by Nanse Mendoza

Chemistry 440 - March 13, 2012

Bibliography

  • Principles of Biochemistry. Robert Horton. 4th Ed.
  • Protein Posttranslational Modifications: The Chemistryof Proteome Diversifications. C.T. Walsh et al. Angew. Chem. Int. Ed. 2005. https://walsh.med.harvard.edu/pubs/PDFs_2/PTM_review.pdf
  • Overview of Post-translational Modifications PTMs. Thermo Scientific. http://www.piercenet.com/browse.cfm?fldID=7CE3FCF5-0DA0-4378-A513-2E35E5E3B49B
  • Protein Covalent Modifications. Harmut Luecke. UC Irvine Department of Biology. http://bass.bio.uci.edu/~hudel/bs99a/lecture26/lecture7_2.html
  • PTMs on H3 variants before chromatin assembly potentiate their final epigenetic state. Loyola, A. et al. Mol Cell. 2006 Oct 20;24(2):309-16. Pub Med.
  • Insulin. Access Science McGraw Hill Science http://www.accessscience.com/popup.aspx?id=347200&name=printEncyclopedia.

Question

In methylation, what is the methyl donor?

S-adenosyl methionine (SAM)

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