J Korean Soc Microbiol.  1998 Apr;33(2):129-137.

Molecular Cloning and Nucleotide Sequence of a Gene Encoding Alcohol Dehydrogenase of Helicobacter pylori

• Rhee KH
• Lee WK
• Cho MJ
• Baik SC
• Jeon YS
• Choi YJ
• Ryu BD
• Song JY
• Lee IG
• Choi SH

Abstract

Partially purified H. pylori ADH was used to determine the amino acid sequence of ADH N- terminus. The sequence of the ADH N-terminus was determined as MRVQSKGF. The genomic library of H. pylori that has been prepared by pTZ19U plasmid vector was screened with the deduced oligonucleotide probes to select the plasmid clone containing the entire ADH gene. The clone pTZ19U/ADH-6 was selected and its EcoRI-BamHI fragment (1.3 kb) was subcloned into pBluescript II K/S vector to determine nucleotide sequence. The length of H. pylori ADH gene was 1,044 bp. Ribosomal binding site was found in the upstream of start codon and rho- independent transcriptional stop signal was observed in the downstream of stop codon. The ADH gene encodes a protein of 348 amino acids, of which the predicted molecular size and pI value were 38.6 kDa and 7.1, respectively. ADH activity of E. coli transformant of pBluescript/ADH is 10-times greater compared to that of non-transformants. When H. pylori ADH gene was disrupted by pBluescript/ADH-KM whose internal region of 1.3 kb DNA fragment containing ADH gene was replaced by KM resistance sequence, the strain lost the ADH activity completely, despite the normal growth of the strain. This demonstrates that ADH gene is not essential for the viability of H. pylori.

MeSH Terms

Alcohol Dehydrogenase*
Amino Acid Sequence
Amino Acids
Base Sequence*
Binding Sites
Clone Cells
Cloning, Molecular*
Codon, Initiator
Codon, Terminator
DNA
Genes, vif*
Genomic Library
Helicobacter pylori*
Helicobacter*
Oligonucleotide Probes
Plasmids
Alcohol Dehydrogenase
Amino Acids
Codon, Initiator
Codon, Terminator
DNA
Oligonucleotide Probes