期刊
BIOCHEMISTRY
卷 56, 期 37, 页码 4951-4961出版社
AMER CHEMICAL SOC
DOI: 10.1021/acs.biochem.7b00499
关键词
-
资金
- National Institutes of Health [R01 DA030604, R01 AT009143, 1R15GM113229-01]
- Loyola University Chicago
- Arthur J. Schmitt Foundation
Potent mechanism-based inactivators can be rationally designed against pyridoxal S'-phosphate (PLP)-dependent drug targets, such as ornithine aminotransferase (OAT) or gamma-amino-butyric acid aminotransferase (GABA-AT). An important challenge, however, is the lack of selectivity toward other PLP-dependent, off: target enzymes, because of similarities in mechanisms of all PLP-dependent aminotransferase reactions. On the basis of complex crystal structures, we investigate the inactivation mechanism of OAT, a hepatocellular carcinoma target, by (1R,3S,4S)-3-amino-4fluorocyclopentane-1-carboxylic acid (FCP), a known inactivator of GABA-AT. A crystal structure of OAT and FCP showed the formation of a ternary adduct. This adduct can be rationalized as occurring via an enamine mechanism of inactivation, similar to that reported for GABA-AT. However, the crystal structure of an off target, PLP-dependent enzyme, aspartate aminotransferase (Asp-AT), in complex with FCP, along with the results of attempted inhibition assays, suggests that FCP is not an inactivator of Asp-AT, but rather an alternate substrate. Turnover of FCP by Asp AT is also supported by high-resolution mass spectrometry. Amid existing difficulties in achieving selectivity of inactivation among a large number of PLP-dependent enzymes, the obtained results provide evidence that a desirable selectivity could be achieved, taking advantage of subtle structural and mechanistic differences between a drug-target enzyme and an off target enzyme, despite their largely similar substrate binding sites and catalytic mechanisms.
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