Biol729 – The human kinome

Protein Kinases The enzymes that catalyze the ATP-dependent phosphorylation of serine, threonine or tyrosine residues in proteins. Essential components of intracellular signalling pathways. When activated, cause phosphorylation and activation/inactivation (through conformational changes) of selected target proteins. This results in a change in cellular activity. The kinome – the full complement of protein kinases encoded by an organism’s genome.

http://pfam.sanger.ac.uk The protein kinase superfamily includes the ser/thr and tyr protein kinases as well as other non-protein kinases.

Eukaryotic protein kinases share a conserved catalytic core common to both serine/threonine and tyrosine protein kinases.

The catalytic subunit of Protein Kinase A (PKA; cyclic AMP-dependent protein kinase) with ATP and a 20 residue segment of a naturally occurring protein kinase inhibitor (pseudo-target sequence: arg-arg-asn-ala-ile). The inhibitor’s pseudo-target sequence is in close proximity to the ATP’s g- phosphate group. In the N-terminal region there is a glycine-rich stretch in the vicinity of a lysine (VAIK) involved in ATP binding. In the central part of the catalytic domain there is a conserved aspartic acid (HRD) important for the catalytic activity of the enzyme. A second aspartic acid (DFG) chelates the Mg2+ ions of ATP. The protein’s main chain is purple with 11 residues that are highly conserved in protein kinases shown in white.

The mouse PKA a-catalytic subunit correlated with crystal structure The mouse PKA a-catalytic subunit correlated with crystal structure. a-Helices are indicated by arrows and b-sheets are boxed. Highly conserved residues are shown in bold, phosphorylation sites are shown as black dots and intron/exon boundaries are indicated by arrows. PKACa SK300 YAMK YRD DFG Active

50 human kinase domains lack at least one of the conserved catalytic residues (lys30, asp125 and asp143) are predicted to be catalytically inactive. It is possible that they use a modified catalytic mechanism that does not require these residues. The Wnk family use lys13 in place of lys30 in ATP binding.

The completion of the human genome project has allowed the identification of almost all human protein kinases. The total (518) is about half that predicted 15 years ago – but it is still a relatively large number ~ 1.7% of all human genes. Most of these protein kinases belong to the superfamily containing the eukaryotic protein kinase catalytic domain. Additionally, 13 atypical protein kinase (aPK) families (40 genes) identified – these contain proteins reported to have biochemical kinase activity, but which lack sequence similarity to the ePK domain.

Dendrogram of 491 ePK domains from 478 human genes.

AGC – cyclic nucleotide regulated protein kinase family, diacylglycerol/phospholipid-dependent protein kinase C family, G-protein-coupled receptor kinases, S6 kinases etc (AGC - protein kinase A, cGMP-dependent protein kinase and protein kinase C ).

Dendrogram of 491 ePK domains from 478 human genes.

CAMK- Ca2+/calmodulin-dependent protein kinases. CKI – casein kinase 1. CMGC – cyclin-dependent kinases, MAP kinases, GSK-3. STE – MAP kinase cascade family members. TK – tyrosine kinases e.g. Src family, Abl family, Tyk2/Jak1 family, EGF receptor family, PDGF receptor family, FGF receptor family, insulin receptor family. TKL – tyrosine kinase-like – a diverse group of families that actually resemble both tyrosine and serine/threonine kinases. e.g. IRAK [interleukin-1 (IL-1) receptor associated kinase]. RGC – receptor guanylate cyclase – similar in domain sequence to the tyrosine kinases.

http://kinweb.ceinge.unina.it/

Graphic showing PKACa – the glycine loop is shown in space-filled blue, lys66, asp160 and asp178 are shown in space-filled green.

Human protein kinase distribution by major groups. Families Subfamilies Protein Kinase Genes Pseudo-genes Novel Kinases AGC 14 21 63 6 7 CAMK 17 33 74 39 10 CK1 3 5 12 2 CMGC 8 24 61 Other 37 83 23 STE 13 47 4 Tyrosine kinase 30 90 Tyrosine kinase-like 43 RGC 1 Human protein kinase distribution by major groups.

All human kinases are found in at least one of the expressed sequence (EST) databases indicating that all are genuine transcribed genes. Many kinases are expressed in low amounts in a restricted distribution. The human kinome also contains many nonfunctional copies of kinase genes (pseudogenes) that are not expressed or encode degenerate, truncated proteins. A few pseudogenes have no obvious human parent but have functional orthologues in rodents and probably indicate the decay of previously functional genes. This PKCι Xq21.3 pseudogene is identical to the PKCι gene sequence except for a point mutation at the stop codon which elongates the ORF by 27 amino acids.

All human kinase genes have been mapped to chromosomal loci All human kinase genes have been mapped to chromosomal loci. Although the overall kinase distribution is similar in density to that of other genes, many pairs of closely related genes from the same families map closely to each other, indicating that they may have arisen through local duplications. Angiogenesis (angio'gen'esis) - the growth of new blood vessels - is an important natural process occurring in the body, both in health and in disease. Chromosomal mapping of the nine human PKC gene loci by FISH (fluorescence in situ hybridization) to metaphase spreads.

Mutations and inappropriate regulation of protein kinases play causal roles in human disease. Agonists and antagonists of these enzymes may be useful for treatment of disease. Comparison of the kinase chromosomal map with disease loci shows that 164 kinases map to amplicons seen in tumours and 80 kinases map to loci implicated in other diseases (e.g. diabetes, hypertension, obesity, osteoarthritis, schizophrenia).

Most protein kinases act in networks and are modulated by autophosphorylation and phosphorylation by other kinases. Other domains within these proteins regulate kinase activity, link to other signalling modules or signal subcellular localization of the protein. 83 additional types of domain, are present in 258 of the 518 kinases. The most common domains mediate interaction with other signalling proteins e.g. 24 kinases contain Src homology 2 (SH2) domains. Other domains link to GTPase signalling, lipid signalling, calcium signalling, target the kinase to the cytoskeleton, or mediate specific interactions with other proteins or RNA.

The PH (pleckstrin homology) domain provides a lipid binding module to direct AKT to PI-3-kinase generated phosphoinositides (PI(3,4,5)P3 and PI(3,4)P2.

Most of the RTKs, 12 receptor serine/threonine kinases and 5 receptor guanylate cyclases have recognizable ligand-binding domains along with signal peptides and transmembrane regions.

Summary A total of 478 human ePKs and 40 aPK genes have been identified in the human genome. Comparison of the human kinase chromosomal map with disease loci shows that kinases map to loci implicated in diseases. Other domains within the protein kinases regulate kinase activity, link to other signalling modules or signal subcellular localization of the protein.

References Manning, G. et al. (2002)Science 298, 1912–1934. The protein kinases of the human genome.