Purification and properties of major alpha-D-mannosidase in the luminal fluid of porcine epididymis.
A lysosomal type alpha-D-mannosidase was successfully purified by DEAE-Sephacel, Red-Amicon and Superdex 200 column chromatographies from porcine cauda epididymal fluid. The purified enzyme consisted of 63 and 51 kDa subunits at equimolar amounts. It cleaved alpha1-2 linked mannosyl residues and less but significantly cleaved alpha1-3 and alpha1-6 linked mannosyl residues in the high-mannose oligosaccharides. The optimal pH to hydrolyze oligosaccharide was in the acidic pH range (pH 3.5 approximately 4.0). Total alpha-D-mannosidase activities in the porcine epididymal fluid increased from proximal to distal caput epididymis, which maintained to cauda epididymis. At least two kinds of alpha-D-mannosidase (lysosomal type enzyme and 135 kDa alpha-D-mannosidase (MAN2B2)) were contained in the porcine epididymal fluid. The activity of the lysosomal type enzyme is much higher than MAN2B2 at the physiological pH. These results suggest that the lysosomal type alpha-D-mannosidase is the predominantly active enzyme in the luminal fluid of porcine epididymis and that it participates in the glycoprotein modification on the sperm surface during epididymal transit.
Institute of Basic Medical Sciences, University of Tsukuba, Tsukuba, Ibaraki 305-8577, Japan.
Y Z
YZ
Jin
F
F
Dacheux
J L
JL
Dacheux
S
S
Bannai
Y
Y
Sugita
N
N
Okamura
D000595:Amino Acid Sequence; D000818:Animals; D001826:Body Fluids / Q000201:enzymology*; D002240:Carbohydrate Sequence; D004822:Epididymis / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D008247:Lysosomes / Q000201:enzymology; D008297:Male; D008361:Mannosidases / Q000737:chemistry / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D013379:Substrate Specificity; D013552:Swine; D043323:alpha-Mannosidase
D008361:Mannosidases; D043323:alpha-Mannosidase