Biology of Reproduction, Vol 24, 691-701, Copyright © 1981 by Society for the Study of Reproduction

Structural Proteins of the Mouse Spermatozoan Tail: An Electrophoretic Analysis

¹ Department of Physiology and Laboratory of Human Reproduction and Reproductive Biology, Harvard Medical School, Boston, Massachusetts 02115

Structural components of the mouse spermatozoan tail were resolved into multiple polypeptide species by using one- and two-dimensional polyacrylamide gel electrophoresis. Spermatozoa were decapitated by brief incubation with either sodium dodecyl sulfate (SDS, 1%) or trypsin (0.1 mg/ml). The SDS- and trypsin-cleaved spermatozoa were subsequently separated into pure head and tail fractions by using sucrose gradient centrifugation and ultracentrifugation in a 40-60% metrizamide gradient, respectively. Analysis of the SDS-insoluble structural tail proteins from trypsin-cleaved and SDS-cleaved spermatozoa by single dimension SDS-polyacrylamide gel electrophoresis revealed approximately 20 protein bands. Following exposure to trypsin several of the high molecular weight components were either reduced in quantity or lost completely. Furthermore, when SDS-insoluble tail proteins were carboxymethylated, the complexity of the polypeptide profile diminished substantially, indicating that some high molecular weight bands represented protein aggregation caused by intermolecular disulfide bonds. Isoelectric focusing (IEF) gels of aminoethylated and carboxymethylated tail proteins also resulted in remarkably different banding patterns. Finally, two-dimensional electrophoresis of the structural tail proteins, reduced and carboxymethylated, enabled definitive resolution of 29 major protein spots.

Note:
ACKNOWLEDGMENTS The authors wish to express their gratitude to Barbara Lewis for preparation of the manuscript and to Steven Borack for assistance in photography. These observations represent partial fulfillment of Honors Thesis by F.M.B. and B.M.M. of Harvard University, Cambridge, MA. The research was funded primarily by NICHHD Grant HD 08270 and in part by NICHHD Centers Grant HD 06645 and Rockefeller Foundation Grant 65040.

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