BIOGENESIS AND SECRETION OF ALKALINE-PHOSPHATASE AND ITS MUTATED FORMS IN ESCHERICHIA-COLI .1. SITE-DIRECTED MUTATIONS IN THE ALKALINE-PHOSPHATASE GENE

Oligonucleotide-directed mutagenesis was used to introduce mutations i nto the E. coli alkaline phosphatase gene, resulting in the following amino acid residue substitutions in the N-terminal part of mature poly peptide chain and in the signal peptide cleavage site: Arg(+1) Ala,Glu (+4) Gln; Glu(+4) Gin; and Ala(-1) Val. Alkaline phosphatase activity was detected in ah E. coli strains transformed with the plasmids conta ining the cloned mutant genes. Besides, an amber mutation was introduc ed into the Arg(+1) position. Active alkaline phosphatase was synthesi zed in E. coli strains containing genes of Ser-, Gln-, Tyr-, Leu-, Ala -, Glu-, Phe-, Gly-, His-, Pro-, and Cys-specific suppressor tRNAs, co nsistent with Arg(+1) substitution by these amino acid residues.