Protein kinase Substrate specificity

Tegge, W., Frank, R., Hofmann, F., and Dostmann, R.G. (1995) Determination of cyclic nucleotide-dependent protein kinase substrate specificity by the use of peptide libraries on cellulose paper. Biochemistry 34, 10569-10577. 

JH Till, RS Annan, SA Carr, WT Miller. Use of synthetic peptide libraries and phosphopeptide-selective mass spectrometry to probe protein kinase substrate specificity. J Biol Chem 269 7423-7428, 1994. 

INVESTIGATION OF THYLAKOID PROTEIN KINASE SUBSTRATE SPECIFICITY USING SYNTHETIC PEPTIDES. 

Investigation of Thylakoid Protein Kinase Substrate Specificity Using Synthetic Peptides 767... 

MARCKS Myristolated, alanine-rich C kinase substrate specific protein kinase C substrate MBP Major basic protein MBSA Methylated bovine serum albumin... 

Proteins from cDNA libraries are used for measurement of kinase substrate specificity and identification of phospholipid-binding proteins. 

Pinna, L.A. Ruzzene, M. (1996) How do protein kinases recognize their substrates Biochvm. Biophys. Acta 1314, 191-225. Advanced review of the factors, including consensus sequences, that give protein kinases their specificity. 

In vitro, with CaATP as a substrate, E. coli dnaK autophosphorylates exclusively at Thr-199 (McCarty and Walker, 1991). It does not auto-phosphorylate when MgATP is used as a substrate. In vivo, dnaK is found to be phosphorylated on serine as well as on threonine residues (Rieul et al., 1987). Under normal growth conditions, phosphorylation is primarily on serine when E. coli is infected with bacteriophage M13, the phosphorylation shifts predominantly to threonine, with minor phosphorylation of serine. The specific target residues of in vivo phosphorylation of dnaK have not yet been determined. The disparity between the in vitro results (autophosphorylation exclusively at Thr-199 an absolute requirement for CaATP) and the in vivo results (phosphorylation on both serine and threonine residues, under conditions in which MgATP would be presumed to be the available intracellular substrate nucleotide) raises the questions of (1) whether the specific autophosphorylation of Thr-199 observed in vitro also occurs in vivo, or whether it may be an artifactual side reaction when the larger Ca ion is substituted for Mg" at the active site of the protein, and (2) whether there is a serine/threonine protein kinase that specifically phosphorylates dnaK in vivo. 

Despite the highly conserved structure of the PTP catalytic domain, PTPs have distinct substrate preferences from one another. For example, PTPIB and TCPTP preferentially dephosphorylate receptor tyrosine kinases and related adaptor molecules, whereas the KIM-family PTPs HePTP, STEP, and PTP-SL dephosphorylate specific MAP kinases. While the presence of distinct non-catalytic domains/motifs facilitate specific localization or binding to substrate proteins, PTP substrate specificity is also dictated by differences within the PTP catalytic domain itself. 

In this method, phosphotransferase activity is used to measure MAPK activity in extracts of smooth muscle tissue. MAPK-specific activity is detected using a synthetic peptide as a substrate in the phosphotransferase reaction. The use of myelin basic protein will result in the detection of all kinases that phos-phorylate this substrate (including MAPK, protein kinase C, and other protein kinases whereas the use of a synthetic peptide, designed as a proline-directed protein kinase substrate, results in the detection of MAPK-specific activity, exclusively (Adam et al., 1995 Clark-Lewis et ah, 1991). 

Fig. 5 Substrate specifity of the purified protein kinase substrate additions (a) purified chromatophores (30/Xg), (b) isolated B875 complexes (30/Xg), (c) histone V-S (15/ig).

Traugh, J.A., Mumby, M., Traut, R.R. Phosphorylation of ribosomal protein by substrate-specific protein kinases from rabbit reticulocytes. Proc. nat. Acad. Sci. (Wash.) 70, 373-376 (1973)... 

In this context, it should be noted that the specific antiviral activity of ganciclovir against HSV (which is more potent than that of acyclovir) can be fully explained by the compound being specifically recognized as substrate by the HSV-encoded TK. For CMV, however, which does not encode a virus-specified TK, the activity of ganciclovir depends on the phosphorylation by a vims-encoded protein kinase, which... 

The roles that Ca + and polyphosphoinositide breakdown products might play in hormone action are presented in Figure 43-6. In this scheme the activated protein kinase C can phosphorylate specific substrates, which then alter physiologic processes. Likewise, the Ca -cahnodulin complex can activate specific kinases. These then modify substrates and thereby alter physiologic responses. 

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