The Aurora A and Aurora B Protein Kinases: A Single Amino Acid Difference Controls Intrinsic Activity and Activation by TPX2

Abstract

The Aurora A and B protein kinases are key players in mitotic control and the etiology of human cancer. Despite the near identity of amino acid sequence in the catalytic domain, monomeric Aurora B is 50 fold lower in activity than monomeric Aurora A, and previous studies have shown that TPX2 binding to the catalytic domain activates Aurora A but not Aurora B. Here we identify G205 in Xenopus Aurora A as a key determinant of both intrinsic activity and regulation by TPX2. Mutation of G205 in Aurora A to N, the equivalent residue in Aurora B, had no effect on autophosphorylation of the T-loop but led to a 10-fold loss of specific activity, whereas mutation of N158 in Aurora B to G caused a 350-fold increase in specific activity. G205 N Aurora A was still activated by TPX2, but protection of pT295 from dephosphorylation by protein phosphatase 1 was abolished. Structural analysis of these effects suggests that the G205 forms a pivot point in the enzyme that results in movement of the N-terminal domain glycine-rich loop closer to the ATP binding site of the enzyme and also moves the C-helix slightly closer to the activation loop. Changes in these positions are comparable to those reported for other protein kinases and demonstrate that phosphorylation of the activation loop alone is not sufficient for enzyme activation. The generation of an activated mutant of Aurora B will be important for studying its role in cell cycle control and tumorigenesis.

A single amino acid change converts Aurora-A into Aurora-B-like kinase in terms of partner specificity and cellular function

“…S2 A). Consistent with previous reports (29,30), the kinase activity of Aurora-A G198N was reduced by 10-fold compared with that of Aurora-A WT (Fig. S3).…”

“…Because the protein sequences of their catalytic domains are very similar, we are interested in understanding how distinct localization and function are determined. Although it has been reported that G198 in human Aurora-A (29) [G205 in Xenopus (30)] is a determinant for its regulation by TPX2 in vitro, the molecular mechanism and the biological effects of the G-N mutation were unknown. Here, we propose that the G-N mutation converts Aurora-A into Aurora-B-like kinase in cell (Fig.…”

Aurora kinases and spindle assembly: Variations on a common theme?

“…Specifically, AurA (G198N) was capable of compensating for the loss of AurB function by forming a complex with INCENP and Survivin and localizing to the inner centromere and the spindle midzone in a manner dependent on its interaction with the components of the CPC. [49][50][51][52] Taken together, these data suggest that the different spatio-temporal distribution and biological functions of AurA and AurB are primarily determined by the nature of their interacting cofactors. If so, can such cofactors, while targeting AurA and AurB to different cellular compartments, still use similar strategies to regulate the kinase activity?…”

VX-680 Inhibits Aurora A and Aurora B Kinase Activity in Human Cells

“…2D), also blocked the phosphorylation of Aurora B on Thr232, which is required for Aurora B phosphotransferase activity. 24,25 We next turned our attention to two putative Aurora A substrates in VX-680-treated cells. The phosphorylation of Aurora A substrates is undetectable in an asynchronous cell population, but Aurora A becomes phosphorylated and activated on Thr288 in mitosis, [26][27][28] when Aurora A phosphorylates the centrosomal protein TACC3/ maskin on Ser558 (human numbering) in both human and Xenopus systems.…”