Single Amino Acid Substitution of Sendai Virus at the Cleavage Site of the Fusion Protein Confers Trypsin Resistance

Abstract

SUMMARYAmino acid sequences of fusion (F) proteins of two trypsin-resistant mutants of Sendai virus, TR-2 and TR-5, were deduced from nucleotide analysis of cDNA encoding the F gene and were compared with that of the trypsin-sensitive wild-type Sendai virus. In both mutants, amino acid substitutions were found at residues 116 (Arg ~ Ile), the cleavage site of the F protein, and 109 (Asn --, Asp). Two trypsinsensitive revertants, TSrev-52 and TSrev-58, derived from TR-5 were both activated by trypsin similarly to the wild-type virus and had a single amino acid reversion from Ile to Arg at residue 116, leaving Asp as before at residue 109. These results indicate that the trypsin sensitivity of Sendal virus can be changed by a single amino acid substitution at the cleavage site of the F protein and a mutation from Arg to Ile is responsible for the acquisition of resistance to trypsin.

Changes in specific cleavability of the Sendai virus fusion protein: implications for pathogenicity in mice

“…The KDe mutants were shown to undergo cleavage activation of the F protein in MDCK cells and were not activated in other cell types, such as LLC-MK2, Madin Derby bovine kidney (MDBK), Vero and L cells (Table 1). In contrast to wt virus, non-active progeny of the KDe mutants produced by a non-permissive cell type, such as LLC-MK2 cells, was not activated by trypsin in vitro but had an increased sensitivity to elastase and, less efficiently, to chymotrypsin, similarly to the elastase activation mutants TR-2, TR-5, pa-c 1 and pa-e2 described previously (Itoh et al, 1987;Scheid & Choppin, 1976;Tashiro & Homma, 1983a). This was shown by multicycle replication experiments in the absence or presence of the different proteases in the culture medium ( Fig.…”

“…On the other hand, protease (elastase) activation mutants, TR-2, TR-5 and pa-cl, have F proteins which are cleavable by chymotrypsin (EC 3.4.21.1) and elastase (EC 3.4.21.36), but are resistant to trypsin and proteases present in various cell types and in chick embryos (Itoh et al, 1987;Hsu et al, 1987;Scheid & Choppin, 1976;Tashiro & Homma, 1983a, b). Infection by the mutants is restricted to a single cycle of replication in mouse lungs, since progeny virus remains non-infectious at this site (Itoh et al, 1990;Hsu et al, 1989;Mochizuki et aL, 1988;Tashiro & Homma, 1983a, 1985Tashiro et al, 1988a).…”

Isolation of an avirulent mutant of Sendai virus with two amino acid mutations from a highly virulent field strain through adaptation to LLC-MK2 cells.

“…cDNA fragments corresponding to each gene of M1 and MVC11 were obtained by screening the cDNA libraries for M1 and MVC11, respectively, using $#P-labelled cDNAs of the SeV Z strain. The positive cDNA fragments were subcloned into M13 mp18\mp19 and sequenced by the dideoxy termination method as described previously (Itoh & Homma, 1987). The nucleotide substitutions in the P and L genes were confirmed by sequencing at least two independent cDNA clones derived from each of M1 and MVC11.…”

Prediction of the virulencies of some enveloped viruses from the structure of the cleavage recognition site of viral glycoproteins essential for infectivity. I. Calculation of interaction energy.

“…Accordingly, the somewhat higher values of V(V) and V(E) than those of avirulent NDV seem to explain the weak virulency of the Z strain described in the Introduction. The HVJ TR2 strain is a mutant isolated after passages of wild-type HVJ in the presence of chymotrypsin (12). This mutant can be activated in vitro by chymotrypsin, but not by trypsin.…”