The gene encoding for a novel hydrolase, cyclic α-maltosyl-(1→6)-maltose [CMM,
cyclo-{→6)-α-D-Glc
p-(1→4)-α-D-Glc
p-(1→6)-α-D-Glc
p-(1→4)-α-D-Glc
p-(1→}] hydrolase (CMMase), was cloned from the genomic library of
Arthrobacter globiformis M6 and designated
cmmF. The gene consisted of 1,353 bp encoding a protein of 450 amino-acids with a calculated molecular mass of 49,344 Da. The deduced amino-acid sequence showed similarities to cyclodextrinase, maltogenic amylase and neopullulanase. On the other hand, the complete sequence of the α-glucosidase gene (
cmmB), which encodes an enzyme involved in the degradation of CMM, revealed that the gene consisted of 1,704 bp encoding a protein of 567 amino-acids with a calculated molecular mass of 63,014 Da. The four conserved regions common in the α-amylase family enzymes were also found in CMMase and α-glucosidase, indicating that these enzymes should be assigned to this family. The DNA sequence of 5,675 bp analyzed in this study contained another four open reading frames (ORFs), designated
cmmC,
cmmD,
cmmE and
cmmG, downstream of
cmmB.
CmmC,
cmmD and
cmmE were expected to encode proteins concerned with incorporation of CMM
via cell membrane.
CmmG was expected to encode a transcriptional regulator protein. CMMase gene, α-glucosidase gene and another four ORFs formed a gene cluster together with 6-α-maltosyltrasferase gene (
cmmA) which encodes a CMM-forming enzyme, namely
cmmABCDEFG. The results of gene analysis suggested that
A.
globiformis M6 has a unique starch utilization pathway
via CMM.
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