COMPLEMENTARY DEOXYRIBONUCLEIC-ACID CLONING AND CHARACTERIZATION OF MSP-10 - THE MOUSE HOMOLOG OF HUMAN ACROSOMAL PROTEIN SP-10

Complementary DNA encoding the putative mouse homologue for human acro somal protein SP-10, a candidate contraceptive vaccinogen, was cloned and sequenced. The entire open reading frame (amino acids 18 to 261) o f the mouse SP-10 (mSP-10),with the exception of the signal peptide (a mino acids 1 to 17), was placed under the influence of inducible T7 RN A polymerase/promoter system to overproduce recombinant protein (re-mS P-10) in Escherichia coli. A six-histidine tag, which was coexpressed at the carboxyl terminus of re-mSP-10, provided the means for purifica tion of re-mSP-10 by immobilized metal chelation affinity chromatograp hy technique. The level of purity of re-mSP-10 thus obtained was deter mined by 2-dimensional gel electrophoresis to be 98%. Immunoblotting w ith monoclonal and polyclonal antibodies previously generated against human or baboon SP-10 showed that mSP-10 shared significant antigenic similarity with its primate counterparts. The position of mSP-10 in th e mouse genome was next mapped through segregation analysis of an inte rspecific backcross panel of 96 animals. Acrv1 (assigned gene symbol f or mSP-10) was localized in the proximal portion of mouse chromosome 9 in a region that exhibits synteny with human 11q23, the region to whi ch ACRV1 (gene symbol for human SP-10) was previously mapped. These ch aracterizations by combined immunological and gene mapping techniques established the cloned mSP-10 to be the mouse homologue of SP-10.