Extracellular Poly(α-L-guluronate)lyase from Corynebacterium sp.: Purification, Characteristics, and Conformational Properties
Extracellular Poly(α-L-guluronate)lyase from Corynebacterium sp.: Purification, Characteristics, and Conformational Properties
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DOI：
10.1023/a:1022534429792
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发表时间：
1998
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影响因子：
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通讯作者：
Y. Matsubara;Ryoko Kawada;K. Iwasaki;T. Oda;T. Muramatsu
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文献类型：
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作者：
Y. Matsubara;Ryoko Kawada;K. Iwasaki;T. Oda;T. Muramatsu
Extracellular alginate lyase was purified from the culture supernatant ofCorynebacteriumsp. isolated from the sewage of a sea tangle processing factory in order to elucidate the structure—function relationship of alginate lyase. The electrophoretically homogeneous enzyme was shown to have a molecular mass of 27 kDa by sodium dodecyl sulfate (SDS)—polyacrylamide gel electrophoresis (PAGE) and by gel filtration, with an isoelectric point of 7.3. The molecular mass from amino acid analysis was 28.644 kDa. The optimalpH and temperature for the enzyme reaction were around 7.0 and 55°C, respectively. Metal compounds such as MnCl2and NiCl2increased the enzyme activity. The enzyme was identified as the endolytic poly(α-L-guluronate)lyase, which was active on poly(α-L-1,4-guluronate) and caused a rapid decrease in the viscosity of alginate solution. Measurement of the far-UV circular dichroic spectrum of the enzyme molecule gave a spectrum with a deep trough at 215nm accompanied by a shallow one at around 237 nm, and with a high peak at 197 nm and a much lower one at 230 nm. This spectrum was most likely to be that of the β-form of the enzyme molecule and resembled poly(β-D-mannuronate)lyase fromTurbo cornutus(wreath shell) and poly(α-L-guluronate)lyase fromVibriosp. (marine bacterium). The near-UV circular dichroic spectrum was characteristic for aromatic amino acid residues. In the presence of 6 M urea, these spectra changed drastically in the near-UV and a little in the far-UV with the disappearance of the enzyme activity. Removal of the denaturant in the enzyme solution by dialysis restored both the activity and inherent circular dichroic spectra. The β-sheets observed in alginate lyases as the major ordered structure seem to be a common conformation for the lyases.
