Abstract
Expression and function of the HSD-3.8 gene encoding a testis-specific protein
Lin,Wen; Zhou,Xuefeng; Zhang,Meilin; Li,Yan; Miao,Shiying; Wang,Linfang; Zong,Shudong; Koide,Samuel S.
Molecular Human Reproduction 7(9): 811-818
Publication date: 2001
The nucleotide sequence of the full length HSD-3.8 cDNA (accessionnumber AF311312), encoding a human sperm component, was determinedto consist of 3818 bp with a reading frame of 2778 bp encodinga deduced polypeptide composed of 926 amino acids. A 0.7 kbfragment containing three immunological epitopes of HSD-3.8cDNA was prepared and used to construct recombinant expressionvectors. The constructs were transformed into E.coli BL-21,and the fusion proteins were expressed, isolated and purified.Using the polyclonal antibodies raised against the purifiedexpressed fusion proteins, positive immunostaining occurredover the surface of the postacrosomal zone of human spermatozoaand of germ cells within the seminiferous epithelium of humantestis. Intense staining of large pachytene primary spermatocytesoccurred. The capacity of the recombinant protein to reducefertility as an immunogen in adult female rats was assessed.Immunized animals were infertile or exhibited marked reductionin their fertility. Analysis of the deduced HSD-3.8 polypeptiderevealed the presence of a tetratricopeptide repeat (TPR) motif,a P-loop sequence that acts as a binding site for ATP/GTP andphosphorylation sites for PKC, CK2 and cAMP/cGMP-dependent proteinkinases. A blot overlay assay with [-P]GTP showed that thepolypeptide encoded by the 0.7 kb fragment of HSD-3.8 is a GTPbinding protein. It was also shown to possess GTPase activityand to be phosphorylated by PKC in vitro. In conclusion, HSD-3.8is a GTP binding protein and its activity may be regulated byphosphorylation.
Get Involved
- Make a contribution to the Population Council
- Sign up to receive e-mail announcements
Connect
