Extracellular metalloprotease gene of Streptomyces cacaoi: structure, nucleotide sequence and characterization of the cloned gene product

Poa Chun Chang, Tai Chih Kuo, Akira Tsugita, Yan Hwa Wu Lee

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

The gene (npr) encoding an extracellular neutral metalloprotease (Npr) from Streptomyces cacaoi YM15 was cloned in Streptomyces lividans using pIJ702 as a vector. The nucleotide (nt) sequence of npr was determined. The deduced open reading frame encoded 550 amino acids (aa)(60 kDa) with a putative signal sequence of 34 aa at the N terminus. High-resolution S1 mapping identified the transcriptional start point at about 132-134 nt upstream from the start codon. The nt sequences at both -10 and -35 regions resemble the consensus sequence of typical Escherichia coli promoters and a fragment containing the promoter was functional in an E. coli promoter probe plasmid. In vitro transcription and translation of the cloned npr sequence revealed a 60-kDa protein product, correlated with the sequence data but not with the size (35 kDa) of the extracellular Npr. The N-terminal as sequence in conjunction with the aa composition analyses on the purified mature Npr led to the conclusion that it was processed from the 60-kDa pre-proenzyme form encoded by npr. The Npr protease contained putative zinc ligand-binding regions and two repeated motifs. Asp-Ser-Gly, similar to the active site residues of the aspartic acid and retroviral proteases.

Original languageEnglish
Pages (from-to)87-95
Number of pages9
JournalGene
Volume88
Issue number1
DOIs
Publication statusPublished - Mar 30 1990
Externally publishedYes

Keywords

  • gene expression
  • in vitro transcription and translation
  • pre-proenzyme
  • Recombinant DNA
  • S1 mapping
  • secretion
  • signal sequence
  • zinc-binding region

ASJC Scopus subject areas

  • Genetics

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