Ack family non-receptor tyrosine kinases are unique with regard to their domain composition and regulatory properties. Human Ack1 (activated Cdc42-associated kinase) is ubiquitously expressed and is activated by signals that include growth factors and integrin-mediated cell adhesion. Stimulation leads to Ack1 autophosphorylation and to phosphorylation of additional residues in the C-terminus. The N-terminal SAM domain is required for full activation. Ack1 exerts some of its effects via protein-protein interactions that are independent of its kinase activity. In the basal state, Ack1 activity is suppressed by an intramolecular interaction between the catalytic domain and the C-terminal region. Inappropriate Ack1 activation and signaling has been implicated in the development, progression, and metastasis of several forms of cancer. Thus, there is increasing interest in Ack1 as a drug target, and studies of the regulatory properties of the enzyme may reveal features that can be exploited in inhibitor design. 1. Introduction The mammalian nonreceptor tyrosine kinases (NRTKs) are divided into ten families: Src, Abl, Jak, Ack, Csk, Fak, Fes, Frk, Tec, and Syk [1]. In addition to their tyrosine kinase catalytic domains, they all contain noncatalytic domains that are important in enzyme regulation and substrate recognition [2]. Like all eukaryotic protein kinases, NRTK catalytic domains have an N-terminal lobe (N-lobe) that contacts ATP, and a larger C-terminal lobe (C-lobe). The activation state of the tyrosine kinases depends on the orientation of an alpha helix (αC) located in the N-lobe. In the active conformation, the αC helix projects inward toward the ATP-binding site [3]. The conformation of a flexible segment in the C-lobe (the activation loop) also has a key role in the regulation of the enzyme activity [4]. The regulatory importance of the phosphorylation of the activation loop varies in the different families of NRTKs. For instance, Src activation is strongly dependent on autophosphorylation [5], while Csk lacks the autophosphorylation site [6]. The contributions of the noncatalytic domains to enzyme regulation are well understood in the case of the Src-family of NRTKS. These enzymes have an N-terminal myristoylation site followed by an SH3 domain, an SH2 domain, and a kinase domain. Two tyrosine residues are important for their activity: Tyr 416 in the activation loop and Tyr 527 in the C-terminal tail. Structural studies showed the spatial arrangement of Src domains in the inactive state, in which the noncatalytic domains mediate intramolecular
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