IDENTIFICATION OF THE ACTIVE-SITE RESIDUES OF THE L-PROTEINASE OF FOOT-AND-MOUTH-DISEASE VIRUS

被引:66
作者
PICCONE, ME
ZELLNER, M
KUMOSINSKI, TF
MASON, PW
GRUBMAN, MJ
机构
[1] USDA ARS,PLUM ISL ANIM DIS CTR,N ATLANTIC AREA,GREENPORT,NY 11944
[2] USDA ARS,EASTERN REG RES CTR,N ATLANTIC AREA,PHILADELPHIA,PA 19118
关键词
D O I
10.1128/JVI.69.8.4950-4956.1995
中图分类号
Q93 [微生物学];
学科分类号
071005 ; 100705 ;
摘要
The foot-and-mouth disease virus (FMDV) leader (L) protein is involved in autocatalytic cleavage at the L/P1 junction and in the cleavage of translation initiation factor p220, a submit of the cap-binding protein complex. It has been suggested that this proteinase has homology to the papain-like family of cysteine proteinases, and from this information, we have investigated the active-site residues by introducing specific mutations into the L gene. Mutations of Cys-23 to Ala or His-120 to Leu resulted in enzymes that lacked cia activity at the L/VP4 cleavage site, trans activity on a truncated L-P1 substrate, and p220 cleavage activity. Mutations of Cys-23 to Ser or His-110 to Leu resulted in enzymes that retained some or all cis activity and had reduced p220 cleavage. These mutations were introduced separately into a full-length FMDV cDNA, and RNA transcripts derived from these cDNAs were translated in a cell-free system and transfected into cells. The C23S mutant inefficiently cleaved at the L/P1 junction and within p1, and virus obtained from transfected cells reverted to wild type. The H110L mutant cleaved the L/P1 junction almost as well as the wild-type enzyme, and virus recovered from transfected cells retained the mutation and displayed wild-type viral protein synthesis and host shut-off kinetics.
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页码:4950 / 4956
页数:7
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