Kinase binding partners

In this technology, the cell or tissue preparation is incubated with the beads to allow specific binding of the kinases, and then the beads are washed extensively under non-denaturing conditions to remove nonspecific proteins. The specifically bound kinases (and kinase binding partners) are eluted under denaturing conditions, subjected to proteolysis to liberate constituent peptides, and the latter are subsequently analyzed and sequenced by mass spectrometry methods. 

PTKs can be subdivided into two large families, receptor tyrosine kinases (RTKs) and non-RTKs. The human genome encodes for a total of 90 tyrosine kinases of which 32 are nonreceptor PTKs that can be placed in 10 subfamilies (Fig. 1). All nonreceptor PTKs share a common kinase domain and usually contain several additional domains that mediate interactions with protein-binding partners, membrane lipids, or DNA (Table 1). These interactions may affect cellular localization and the activation status of the kinase or attract substrate proteins for phosphorylation reactions. 

The Crk protein was first discovered as the transforming principle of the retroviruses CTIO and ASV-1. Abl tyrosine kinase is under discussion as a binding partner of the SH3 domain of Crk (Feller et al., 1994). Possible binding partners of the SH2 domain have been described but their physiological function is unclear. 

In some cases, the domain-binding partner is internal. Phosphorylation of some protein kinases inhibits their activity by favoring the interaction of an SH2 domain with a (P)-Tyr in another domain of the same enzyme. For example, the soluble protein Tyr kinase Src, when phosphorylated on a critical Tyr residue, is rendered inactive as an SH2 domain needed to bind to the substrate protein instead binds to an internal (P)-Tyr... 

Q331S6] 2) Casein kinase-1 binding partner (CK1 BP) 3) Stem cell factor apoptosis response protein-1 (SCF ARP1), isoform a (55861) [15881]... 

Dysbindin-2B has been identified as both stem cell factor apoptosis response protein 1 (SCF ARP1) by Lucas et al. (2005) and as a casein kinase 1 binding partner (CK1BP) by Yin et al. (2006). SCF ARP1 is expressed in bone marrow-derived myelomonocytic stem cells upon induction of apoptosis by withdrawal of stem cell growth factor SCF (Lucas et al., 2005). Programmed cell death via apoptosis controls stem cell numbers during hematopoietic cell development (Domen 2001). Since apoptosis is similarly important in development of neurons (Kuan et al., 2000 Buss and Oppenheim, 2004), dysbindin-2B may play a role in nervous system development (see also Section 2.2.33.1). 

Further discussion of the dysbindin family is limited to dysbindin-1 since little is known about other family members apart from the fact that dysbindin-2B is a casein kinase 1 binding partner and a stem cell factor apoptosis response protein. In contrast, much has been learned about dysbindin-1 since its discovery was reported by Benson et al. (2001). It is the only member of the dysbindin family known to exist in invertebrates, specifically the fruit fly, and may thus date back 600 million years. Unlike all other dysbindin paralogs, it contains a coiled coil domain (CCD) allowing extensive interactions with other proteins. A leucine zipper motif in the CCD changed in the course of evolution in a manner permitting more durable interactions with binding partners. 

Yin H, Laguna KA, Li G, Kuret J. 2006. Dysbindin structural homologue CK1BP is an isoform-selective binding partner of human casein kinase-1. Biochemistry 45 5297-308. 

In summary, the KinaTor technology appears to be extremely potent for potentially revealing all binding partners of biologically active kinase inhibitors. Transmembrane domain proteins bind, as well as lipid kinases and other nucleotidebinding proteins, such as heat-shock proteins and oxidoreductases (unpublished data). So far, there are no limitations. 

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