DIFFERENT REQUIREMENTS FOR PRODUCTIVE INTERACTION BETWEEN THE ACTIVE-SITE OF HIV-1 PROTEINASE AND SUBSTRATES CONTAINING -HYDROPHOBIC-ASTERISK HYDROPHOBIC- OR -AROMATIC-ASTERISK PRO- CLEAVAGE SITES
DIFFERENT REQUIREMENTS FOR PRODUCTIVE INTERACTION BETWEEN THE ACTIVE-SITE OF HIV-1 PROTEINASE AND SUBSTRATES CONTAINING -HYDROPHOBIC-ASTERISK HYDROPHOBIC- OR -AROMATIC-ASTERISK PRO- CLEAVAGE SITES
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DOI：
10.1021/bi00137a015
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发表时间：
1992-06-09
期刊：
影响因子：
2.9
通讯作者：
KAY, J
中科院分区：
文献类型：
--
作者：
GRIFFITHS, JT;PHYLIP, LH;KAY, J
已经研究了 HIV-1 蛋白酶催化切割含有两种不同类型连接（-疏水*疏水-或-芳香*Pro-）的寡肽的序列要求。对于第一种类型的连接（-疏水*疏水-），根据最近对自然切割位点的统计分析，发现 P2 和 P2' 位置的最佳残基分别是 Val 和 Glu [Poorman, R. A., Tomasselli, A. G., Heinrikson, R. L., & Kezdy, F. J. (1991) J. Biol.化学。 266、14554-14561]。对于-aromatic*Pro-类型的连接，在此处研究的特定序列背景中，再次观察到P2'位置中的Glu值。假定的酶-底物复合物的分子模型提供了对具有序列 -Val-Tyr*Pro- 的肽所观察到的低效裂解的解释。还记录了 P5 位点的氨基酸对切割率的显着影响。而疏水*疏水型的一个序列中 P5 位置的赖氨酸会产生一种非常有效地裂解的肽（对于 Lys-Ala-Arg-Val-Nle*p-硝基苯基-丙氨酸-P2'-Ala-Nle-NH2，对于 P2' = Glu、Gln、Ile、Val 或 Ala，k(cat) > 15 s-1），对于-芳香族*Pro-型，P5 残基可以对裂解率产生积极或消极的影响。这些结果再次根据分子模型进行了解释。我们认为活性位点裂缝外围的底物序列的相互作用可能影响酶-底物对的匹配，从而控制催化效率。因此，HIV-1 PR 选择性且有效地切割各种完全不同序列的能力可能部分源于距实际可断裂肽键较远距离的广泛相互作用以及酶的多肽结构的几个关键环的固有灵活性。
The sequence requirements for HIV-1 proteinase catalyzed cleavage of oligopeptides containing two distinct types of junctions (-hydrophobic*hydrophobic- or -aromatic*Pro-) has been investigated. For the first type of junction (-hydrophobic*hydrophobic-) the optimal residues in the P2 and P2' positions were found to be Val and Glu, respectively, in accord with recent statistical analysis of natural cleavage sites [Poorman, R. A., Tomasselli, A. G., Heinrikson, R. L., & Kezdy, F. J. (1991) J. Biol. Chem. 266, 14554-14561]. For the -aromatic*Pro- type of junction, in the specific sequence context studied here, the value of Glu in the P2' position was again observed. An explanation for the inefficient cleavage observed for peptides with the sequence -Val-Tyr*Pro- has been provided from molecular modeling of the putative enzyme-substrate complex. A significant effect upon cleavage rates due to the amino acid in the P5 position has also been documented. While lysine in the P5 position in one sequence of the -hydrophobic*hydrophobic-type produces a peptide cleaved very efficiently (k(cat) > 15 s-1 for Lys-Ala-Arg-Val-Nle*p-nitrophenyl-alanine-P2'-Ala-Nle-NH2, for P2' = Glu, Gln, Ile, Val, or Ala), for substrates of the -aromatic*Pro- type, the P5 residue can exert either a positive or negative effect on cleavage rates. These results have again been interpreted in light of molecular modeling. We suggest that interaction of the substrate sequence on the periphery of the active site cleft may influence the match of the enzyme-substrate pair and, hence, control the efficiency of catalysis. Thus, ability of HIV-1 PR to selectively and efficiently cleave a variety of totally different sequences may be derived, in part, from extensive interactions at long distances from the actual scissile peptide bond and the inherent flexibility of several key loops of polypeptide structure of the enzyme.