Purification and characterization of a beta-galactosidase from peach (Prunus persica).
A beta-galactosidase (EC 3.2.1.23) from peach (Prunus persica cv Mibackdo) was purified and characterized. The purified peach beta-galactosidase was 42 kDa in molecular mass and showed high enzyme activity against a the beta-galactosidase substrate, rho-nitrophenyl-beta-D-galactopyranoside. The Km and Vmax values of the enzyme activity of the peach beta-galactosidase were 5.16 and 0.19 mM for rho-nitrophenyl-beta-D-galactopyranoside mM/h, respectively. The optimum pH of the enzyme activity was pH 3.0, but it was relatively stable from pH 3.0-10.0. The temperature optimum was 50 degrees C. The enzyme activities were not improved in the buffers that contained Ca2+, Cu2+, Zn2+, and Mg2+, which indicates that the purified peach beta-galactosidase did not require these cations as co-factors. However, the enzyme was completely inhibited by Hg2+. The purified protein was cross-reacted with an antibody against the persimmon fruit beta-galactosidase. A further comparison of the N-terminal amino acid sequence of the purified protein showed high homologies to those of beta-galactosidase in apple (87%), persimmon (80%), and tomato (87%). Therefore, enzymatic, immunological, and molecular evidences in this study indicate that the purified 42-kDa protein is a peach beta-galactosidase.
Lee
Dong Hoon
DH
Department of Horticulture, Yeungnam University, Gyeongsan 712-749, Korea.
Kang
Sang-Gu
SG
Suh
Sang-Gon
SG
Byun
Jae Kyun
JK
D000595:Amino Acid Sequence; D015153:Blotting, Western; D002412:Cations / Q000494:pharmacology; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003429:Cross Reactions; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010940:Plant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D027861:Prunus / Q000201:enzymology* / Q000235:genetics / Q000276:immunology; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D013379:Substrate Specificity; D013696:Temperature; D001616:beta-Galactosidase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism
D002412:Cations; D004791:Enzyme Inhibitors; D010940:Plant Proteins; D001616:beta-Galactosidase