Signal and membrane anchor functions overlap in the type II membrane protein I gamma CAT.

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Abstract
I.gamma.CAT is a hybrid protein that inserts into the membrane of the endoplasmic reticulum as a type II membrane protein. These proteins span the membrane once and expose the NH2-terminal end on the cytoplasmic side and the COOH terminus on the exoplasmic side. I.gamma.CAT has a single hydrophobic segment of 30 amino acid residues that functions as a signal for membrane insertion and anchoring. The singgn-anchor region in I.gamma.CAT was analyzed by deletion mutagensis from its COOH-terminal end (.DELTA.C mutants). The results show that the 13 amino acid residues on the amino-teminal side of the hydrophobic segment are not sufficient for membrane insertion and translocation. Mutant proteins with at least 16 of the hydrophobic residues are inserted into the membrane, glycosylated, and partially proteolytically processed by a microsomal protease (signal peptidase). The degree of processing varies between different .DELTA.C mutants. Mutant proteins retaining 20 or more of the hydrophobic amino acid residues can span the membrane like the parent I.gamma.CAT protein and are not proteolytically processed. Our data suggest that in type II membrane protein I.gamma.CAT, the signals for membrane insertion and anchoring are overlapping and that hydrophilic amino acid residues at the COOH-terminal end of the hydrophobic segment can influence cleavage by signal peptidase. From this and previous work, we conclude that the function of the signal-anchor sequence in I.gamma.CAT is determined by three segments: a positively charged NH2 termimus, a hydrophobic core of at least 16 amino acid residues, and the COOH-terminal flanking hydrophilic segment.