1887

Abstract

Summary: The gene () for the highly thermostable neutral protease of MK232 was cloned in using pTB53 as a vector. The nucleotide sequence of and its flanking regions was determined. The DNA sequence revealed only one large open reading frame, composed of 1656 base pairs and 552 amino acid residues. A Shine-Dalgarno (SD) sequence was found 12 bases upstream from the translation start site (ATG). A possible promoter sequence (TTTTCC for the — 35 region and TATTGT for the — 10 region), which was nearly identical to the promoter for another thermostable neutral protease gene, , was also found about 40 bases upsteam of the SD sequence. The deduced amino acid sequence contained a signal sequence in its amino-terminal region. The sequence of the first five amino acids of purified extracellular protease completely matched residues 237-241 of the open reading frame. This suggests that the enzyme is translated as a large polypeptide containing a pre-pro structure as is known for other neutral proteases. The amino acid sequence of the extracellular form of this protease (316 amino acids, molecular mass 34266 Da) was identical to that of the thermostable neutral protease (thermolysin) from except for two amino acid substitutions (Asp37 to Asn37 and Glu119 to Gln119). The G + C content of the coding region of was 42 mol%, while that of the third letter of the codons was lower (36 mol%). This extremely low content is an exceptional case for genes from thermophiles. When the protease genes, and , were cloned on pTB53 in , the expression of was about 20 times higher than that of The reason for the difference between the two systems is discussed.

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/content/journal/micro/10.1099/00221287-134-7-1883
1988-07-01
2019-10-22
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http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-134-7-1883
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