alpha-Amylase gene of thermophilic Streptomyces sp. TO1: nucleotide sequence, transcriptional and amino acid sequence analysis.
The nucleotide sequence of a 1860-bp region encoding a thermostable alpha-amylase of Streptomyces sp. TO1 was determined. Frame analysis revealed the presence of a 1359-bp long open reading frame (amy TO1) encoding a 453 amino acid protein with a deduced M(r) of 49 kDa. Northern blot analysis revealed that amy TO1 gene was expressed as approximately 1.5-kbp monocistronic transcript in both SL1326/pLM1 and Streptomyces sp. TO1 strains. Primer extension experiments indicated that the transcriptional start site lies 30 bp upstream of the ATG start codon, and allowed the identification of -35 (TTGCTG) and -10 (TACGCG) eubacterial-like promoter sequences. Amy TO1 exhibits strong amino acid identities with those from other Streptomyces species with a maximum of 78% with S. thermoviolaceus alpha-amylase. Nevertheless, subtle amino acid changes such as the substitution of four conserved residues found at similar positions in other Streptomyces alpha-amylases by proline residues, and the substitution of three conserved hydrophilic amino acids by hydrophobic ones in Amy TO1 might account for the thermostable properties of Amy TO1.
Mellouli
L
L
Centre de Biotechnologie de Sfax, Tunisie.
Ghorbel
R
R
Virolle
M J
MJ
Bejar
S
S
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D015152:Blotting, Northern; D017931:DNA Primers; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D006358:Hot Temperature; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000032:analysis; D017422:Sequence Analysis, DNA; D013302:Streptomyces / Q000201:enzymology / Q000235:genetics*; D014158:Transcription, Genetic / Q000502:physiology*; D000516:alpha-Amylases / Q000737:chemistry* / Q000235:genetics*
D001426:Bacterial Proteins; D017931:DNA Primers; D012333:RNA, Messenger; D000516:alpha-Amylases