Lazic Ana, Goetz David H, Nomura Anson M, Marnett Alan B, Craik Charles S
Department of Pharmaceutical Chemistry, University of California, San Francisco, CA 94158-2517, USA.
J Mol Biol. 2007 Nov 2;373(4):913-23. doi: 10.1016/j.jmb.2007.07.073. Epub 2007 Aug 16.
The herpesvirus proteases are an example in which allosteric regulation of an enzyme activity is achieved through the formation of quaternary structure. Here, we report a 1.7 A resolution structure of Kaposi's sarcoma-associated herpesvirus protease in complex with a hexapeptide transition state analogue that stabilizes the dimeric state of the enzyme. Extended substrate binding sites are induced upon peptide binding. In particular, 104 A2 of surface are buried in the newly formed S4 pocket when tyrosine binds at this site. The peptide inhibitor also induces a rearrangement of residues that stabilizes the oxyanion hole and the dimer interface. Concomitant with the structural changes, an increase in catalytic efficiency of the enzyme results upon extended substrate binding. A nearly 20-fold increase in kcat/KM results upon extending the peptide substrate from a tetrapeptide to a hexapeptide exclusively due to a KM effect. This suggests that the mechanism by which herpesvirus proteases achieve their high specificity is by using extended substrates to modulate both the structure and activity of the enzyme.
疱疹病毒蛋白酶是一个通过形成四级结构实现酶活性变构调节的例子。在此，我们报道了卡波西肉瘤相关疱疹病毒蛋白酶与一种六肽过渡态类似物复合物的1.7埃分辨率结构，该类似物可稳定酶的二聚体状态。肽结合后会诱导出延伸的底物结合位点。特别是，当酪氨酸结合在该位点时，表面的104 Ų被埋入新形成的S4口袋中。肽抑制剂还会诱导残基重排，从而稳定氧负离子洞和二聚体界面。伴随着结构变化，延伸底物结合后酶的催化效率会提高。仅由于KM效应，将肽底物从四肽延伸至六肽时，kcat/KM增加近20倍。这表明疱疹病毒蛋白酶实现其高特异性的机制是通过使用延伸底物来调节酶结构和活性。
