The isolation from human seminal plasma of a new form of soluble 5'-nucleotidase.
Soluble broad spectrum 5'-nucleotidase from human seminal plasma was purified to homogeneity by a combination of (NH4)2SO4 precipitation, affinity chromatography, and gel filtration. The pure enzyme had a specific activity of 4800 nmol min-1 mg-1. SDS-PAGE of purified enzyme preparation revealed a single polypeptide band of 53 kDa and a tetrameric structure of 203 kDa was proposed for the native enzyme. This form had modest preference for AMP as substrate; Mg2+ and Mn2+ were activators of the enzyme although its activity was not absolutely dependent on the presence of these exogenous bivalent cations. The enzyme, recovered in the nonsedimentable fraction of human seminal plasma, had a pH optimum of 7.5; ATP and ADP were inhibitors of mixed type, Pi was a potent inhibitor at nonphysiological concentrations, and Con A and adenosine 5-[alpha, beta-methylene]diphosphate had no effect on the enzyme activity. The enzyme described here therefore has some unique properties between truly cytoplasmic and membrane-bound derived forms.
Minelli
A
A
Dipartimento di Biologia Cellulare e Molecolare, Università degli Studi di Perugia, Perugia, 06123, Italy.
Moroni
M
M
Monacelli
N
N
Mezzasoma
I
I
D000241:Adenosine / Q000096:biosynthesis; D000244:Adenosine Diphosphate / Q000378:metabolism / Q000494:pharmacology; D000249:Adenosine Monophosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism / Q000494:pharmacology; D004795:Enzyme Stability; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000302:isolation & purification; D008297:Male; D008670:Metals / Q000494:pharmacology; D008970:Molecular Weight; D009708:Nucleotidases / Q000737:chemistry* / Q000187:drug effects / Q000302:isolation & purification*; D010710:Phosphates / Q000378:metabolism / Q000494:pharmacology; D012661:Semen / Q000737:chemistry / Q000201:enzymology*; D012995:Solubility; D013379:Substrate Specificity; D013696:Temperature
D007527:Isoenzymes; D008670:Metals; D010710:Phosphates; D000249:Adenosine Monophosphate; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D009708:Nucleotidases; C030287:nucleotidase; D000241:Adenosine