Deletions into an NH2-terminal hydrophobic domain result in secretion of rotavirus VP7, a resident endoplasmic reticulum membrane glycoprotein

Bam HI and the 387-bp 5'-terminal fragment of the VP7 gene ( Fig. 1 ) was subcloned into the Barn HI site of pBR322 to generate pBR9B ( Fig. 1) 1 ) and a fragment from the same plasmid, which extended through the VP7 gene, counterclockwise from the EcoR I site to the Cla I site near the 5'-end of the gene (7). The fourth fragment which permitted the vector to circularize was an Xho 1/Nco I fragment of 53 bases derived from a pBR9B deletion mutant ( Fig. 1 ) , where the Bal 31 digestion went precisely to the first ATG codon. This construction deleted the first eight amino acids of VP7 which are conserved between human, simian, and bovine rotaviruses, and substituted Met-Ala-Met such that the final NH2-terminal sequence now reads as Met-Ala-Met-Val-Leu-Thr .......