POINT MUTATION IN AVIAN-SARCOMA LEUKEMIA-VIRUS PROTEASE WHICH INCREASES ITS ACTIVITY BUT IMPAIRS INFECTIOUS VIRUS PRODUCTION

被引:10
作者
ARAD, G
CHOREV, M
SHTORCH, A
GOLDBLUM, A
KOTLER, M
机构
[1] HEBREW UNIV JERUSALEM, HADASSAH MED SCH, DEPT MOLEC GENET, IL-91010 JERUSALEM, ISRAEL
[2] HEBREW UNIV JERUSALEM, HADASSAH MED SCH, DEPT PHARMACEUT CHEM, IL-91010 JERUSALEM, ISRAEL
关键词
D O I
10.1099/0022-1317-76-8-1917
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
The retrovirus protease (PR), an aspartic PR, is composed of two identical subunits, each containing a conserved tripeptide sequence present at the active site of the enzyme. Asp-Ser-Gly is found in avian sarcoma leukaemia viruses (ASLV) and Asp-Thr-Gly in mammalian oncoretroviruses. We have mutated the conserved sequence at the active site of ASLV PR by converting the Ser Gly residues to Thr and Ala, respectively. Replacement of Gly with Ala yielded an ASLV PR devoid of proteolytic activity. The Ser to Thr conversion did not alter the substrate specificity of the enzyme. Both wild-type and mutated PRs correctly cleaved viral precursors expressed in bacterial cells, as well as synthetic peptides homologous to ASLV and human immunodeficiency virus type 1 cleavage sites. Bacterially produced ASLV PR with Thr instead of Ser had increased enzymatic activity, as shown by hydrolysis of synthetic peptides. However, this mutation reduced the production of reverse transcriptase-containing particles and infectious virus following transfection of permissive cells with virus DNA.
引用
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页码:1917 / 1925
页数:9
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