The PH-20 protein in human spermatozoa.
PH-20 is a sperm plasma-membrane protein that has been shown to have hyaluronidase activity in several mammalian species including nonhuman primates. In this investigation, the PH-20 protein was characterized in noncapacitated human sperm and in capacitated human sperm. Two forms of PH-20 were observed in immunoblots of sodium dodecylsulfate polyacrylamide-gel electrophoresis (SDS PAGE) using a polyclonal antibody to recombinant PH-20: a major band of 64 kDa appeared in noncapacitated and capacitated sperm extracts and a 53-kDa band that appeared only in the acrosome-reaction supernatant of acrosome-reacted sperm. Using hyaluronic acid substrate gel analysis, we demonstrated that noncapacitated sperm extracts, capacitated sperm extracts, and the acrosome-reaction supernatant had hyaluronidase activity at neutral pH (pH 7) and acid pH (pH 4). The 64-kDa form in all samples had hyaluronidase activity at both neutral and acid pH, but the 53-kDa form was only active at acid pH. Total hyaluronidase activity, as measured by a microplate assay, was higher at pH 7 than at pH 4. Very low hyaluronidase activity was detected in the acrosome-reaction supernatant. Transmission electron microscopy and immunogold labeling showed that PH-20 of acrosome-intact human sperm was located on the plasma membrane over the entire head but not on the sperm midpiece and tail. After the acrosome reaction, PH-20 was also located on the inner acrosomal membrane. The biochemical characteristics and the ultrastructural localization of PH-20 in human sperm suggest that this protein is the human sperm hyaluronidase and, therefore, has an important function during fertilization.
Sabeur
K
K
Department of Obstetrics and Gynecology, University of California, Davis 95616, USA.
Cherr
G N
GN
Yudin
A I
AI
Primakoff
P
P
Li
M W
MW
Overstreet
J W
JW
D000177:Acrosome / Q000502:physiology / Q000648:ultrastructure; D015153:Blotting, Western; D015815:Cell Adhesion Molecules / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006801:Humans; D006821:Hyaluronoglucosaminidase / Q000032:analysis; D008297:Male; D008565:Membrane Proteins / Q000032:analysis; D013075:Sperm Capacitation; D013084:Sperm-Ovum Interactions; D013094:Spermatozoa / Q000737:chemistry* / Q000502:physiology*
D015815:Cell Adhesion Molecules; D008565:Membrane Proteins; D006821:Hyaluronoglucosaminidase; C066852:hyaluronidase PH-20