PURIFICATION AND SOME CHARACTERISTICS OF PECTATE LYASE FROM PSEUDOMONAS FLUORESCENS GK-5

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Abstract
Pseudomonas fluorescens GK‐5 produced up to 35 U per ml of a single extracellular pectate lyase in a gluconate‐yeast extract medium. The enzyme was purified by chromatography on CM Bio‐Gel A at pH 7.0, and on crosslinked pectate, first at pH 7.0 and then at pH 9.0. The pure enzyme (specific activity 956 U per mg protein) has a molecular weight of 42300 and an isoelectric point of 10.3. Its amino acid composition was determined. The pectate lyase is an endo enzyme, requiring calcium ions for its activity. The enzyme is maximally active at pH 9.4 and ionic strength 0.01. It has a Km value of 0.10 mg polygalacturonic acid per ml and a Vmax of 1.3 millimoles of unsaturated products released per min per mg of enzyme protein. The organism produces soft rot in potato. The enzyme macerates potato tissue optimally at about pH 8. Cell‐bound pectate lyase was also found. Molecular and kinetic properties of the cell‐bound enzyme are identical to those of the extracellular enzyme.
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