PROTEIN PHOSPHORYLATION

By:Sachin kumarM.Pharm. (Pharmacology)Deptt. of Pharma.

SciencesM.D.U. Rohtak, 124001

CONTENTSIntroductionProtein phosphorylation sites.Mechanism of phosphorylation.Function of phosphorylation.Role of phosphorylation in human

health & disease. Disease caused by abnormal

phosphorylation.Methods of detection of protein

phophorylation.Recent studies References

PROTEIN Protein are biomolecules cosisting of one

or more long chain of amino acid residue.

PROTEIN PHOSPHORYLATIONProtein phosphorylation was first reported

in 1906 by Phoebus Levene. He identified phosphate in protein vitellin.

It is a post-translational modification of protein in which an amino acid residue is phosphorylated by a protein kinase by the addition of covalently bound phosphate group.

It alters the structural conformation of a protein , causing it to become activated, deactivated or modifying its function.

De-phosphorylation-The de-phosphorylation of phosphoprotein is

carried out by another group of enzyme called protein phosphatase.

Protein phosphorylation sitesMost commonly phosphorylated amino acids

are serine, threonine and tyrosine.There are thousand of different kind of

protein in a cell, hence there are thousand of distinct phosphorylation sites in a given cell.

 One study indicates that 30% of proteins in the  human  genome can be phosphorylated, and  abnormal phosphorylation is now recognized as a cause of  human  disease.

Mechanism of p.phophorylation Amino acids serine, threonine & tyrosine

have a neuclophilic (-OH) group that attacks the terminal phosphate group on the universal phophoryl donor ATP, resulting in the transfer of phosphate group to the amino acid side chain.

Functions of phosphorylation

MOLECULAR CELLULAR

• Subcellular location • Signal transduction

• Enzyme activity • Cell division

• Ligand binding • Differentiation

• DNA binding • Virulence

• Prot.-prot. Interaction • Cell transformation

• Protein degradation

Role of phosphorylation in human health & disease

•DISEASE •KINASE/PHOSPHOTASE

•Diabetes • Insulin receptor kinase•Colon, breast & other sporadiac cancer

•PI-3 kinase

• Herediatry Parkinson disease

• LRRK2

• X-Linked myotubular myopathy

• MTMI tyrosine phosphotase

• Gordon hypertension syndrome

•WNK1 & WNK4

• Non-hodgkins lymphona •AIK kinase

First protein kinase & phosphotase inhibitor to be used clinically……

Cyclosporin Cyclosporin- Cyclophillin

NF-AT Calcineurin (PP2B) NF-AT

Interlukin -2

T-cell proliferation

cause Tumour . (cancer)

P

Protein kinase inhibitor as drug to treat cancer… Interaction of growth factor (EGF & PDGF) with their

receptor. Activate tyrosin kinase STI571 GTP-Ras Abelson

STI571 tyrosine

kinase Raf PD98059,UO126 MKK1 (MAPK kinase1) MAPKs

--

--

--

CDKs

Cell proliferation

• Over expression and mutation of receptor and other protein tyrosin kinase such as Abelson tyrosine kinase cause cell transformation and cancer.

• The EGF receptor kinase is over expressed in many cancer such as lungs and breast cancer.

ROSCOVITINEFLAVOPIRIDOLINDIRUBIN

--

Protein kinase inhibitors as drug to treat chronic inflammatory disease….

LPS (Lipopolysaccharide)

SAPK2a/p38 SB203580

MAPKAP-K2

TNF (Tumour necrosis factor)

SB203580 SAPK2a/p38 IL-6

--

--

GSK3 inhibitors as drug to treat diabetes…. Insulin

Insulin receptor Tyrosine kinase

IRS PI3 PIP2 PIP3 PDK1

It phosphorylate GSK3 & Protein kinase B Inhibit its dephophorylation

GSK3 (Inhibited)

GS elF2B

Glycogen Protein Synthesis Synthesis• Type –II or NIDDM, which accounts for 90%

of all cases of this disease is characterized by insulin resistance .

• Insulin induced inhibition of GSK3 and treat diabetes.

Disease caused by abnormal phosphorylationAbnormal phosphorylation cause no. of

disease such as Alzheimer‘s Disease, Parkison Disease & other degenerative disorders.

In Alzheimer‘s Disease hyperphosphorylation of Tau protein leads to abnormal aggregation into fibrillary tangles inside the neuron & cause cell death.

In Parkinson Disease phosphorylation of Ser¹²⁹ is associated with the aggregation of protein & further damage to the nervous system.

Methods of detection of protein phophorylation…..

Western blottingELISA (enzyme linked immunosorbent assay)Quantitative mass spectrometryIntracellular flow cytometry

RECENT STUDY………. In recent study it is found that circadian

clock exerts grater control via protein phophorylation which modulate protein activity.

Previous study indicated that protein phosphorylation play a important role in controlling the the circadian clock molecular mechanism.

Scientists did not know how big? Maria Robels, a choronobiologist at the

Max planck institutes of biochemistry in Planegg Germany, used high resolution mass spectrometry to detect flcutuation in the amount of phosphorylated protein in mouse liver cell over a 24hr period.

About 40% of phophorylated protein in mouse liver displayed distinct circadian oscillation.

References HRV A. Synthesis of amino acids and proteins vol.7.In

MTP International Review of Sciences. Biochemistry Section. University Park Press, Baltimore, Md., 1975.

Ptacek J, Synder M, Global analysis of protein phophorylation.2006, PMID: 16908088.

https://www.rndsystems.com/resources/articles/methods detecting protein phosphorylation.

P.Cohen, The role of protein phosphorylation in human health and disease, the FEBS journal, 2001, vvol.268. http://onlinelibrary.wiley.com/doi/10.1046/j.0014 2956.2001.02473.

http://journal.frontiersin.org/article/10.3389/fmicb.2016.00184.

http://blog.pnas.org/2017/02/journal club phosphorylation has surprisingly big role in control of circadian clocks.

Thomas B,M. Flint, Human molecular genetics (2007) 16 (R2): R183-R194. DOI: https://doi.org/10.1093/hmg/ddm159 Published: 15 October 2007

http://neurosciencenews.com/alzheimers kinase p38y 5549.

Kolarova M,Bartos J,Ricny J,Ripova D, Structure and Pathology of Tau Protein in Alzheimer Disease , International Journal of Alzheimer's Disease, volume  2012, Article ID:731526.