Mitsuwo Hara

The chymotrypsin-like activity of human prostate-specific antigen, γ-seminoprotein

γ‐Seminoprotein (γ‐Sm) is a human prostate‐specific antigen and a serine protease judging from the complete amino acid sequence which shows extensive homology with the kallikrein family. The enzymatic activity of γ‐Sm was defined as a chymotypsin‐like activity using reduced and S‐3‐(trimethylated amino)propylated lysozyme and insulin‐oxidized A and B chains as substrates. The ‐Leu Ser‐ peptide bond of lysozyme was rapidly hydrolyzed by γ‐Sm. γ‐Sm also hydrolyzed the ‐Phe Glu‐ of lysozyme and the ‐Leu Cys(SO3H) ‐of insulin B chain. Insulin A chain and arginyl‐ or lysyl‐linkage of these proteins were not hydrolyzed by γ‐Sm at all.

The amino acid sequence of human β-microseminoprotein

Abstract The complete amino acid sequence of β-microseminoprotein of human seminal plasma was determined by automated Edman degradation of the protein and peptides which were obtained by enzymatic cleavage with trypsin, chymotrypsin and Staphylococcus aureus V8 proteinase. The carboxyl-terminal sequence of the protein was established with the aid of carboxypeptidase A. The amino acid sequence of this protein proved to be as follows: Thus, β-microseminoprotein consisting of 93 amino acid residues has a molecular mass of 10 652 Da. The linear structure of this protein represents the first complete amino acid sequence of a sperm-coating protein specific to human seminal plasma.

Occurrence of the Y determinant on the N-glycosidic carbohydrate units of human gamma-seminoprotein

Abstract 500-MHz 1 H-NMR spectroscopy of the oligosaccharides derived from γ-seminoprotein, a human seminal plasma glycoprotein, revealed considerable microheterogeneity both with respect to the degree of branching and with regard to the peripheral sugars. Although the protein possesses only one N-glycosylation site, di-, tri- and tetra-antennary glycans in a ratio of 40:15:45 were found to be present. Moreover, certain branches of the tri- and tetra-antennas contain one or two Fuc residues which form part of the SSEA-1 and Y determinants. It should be noted that this is the first report that describes the occurrence and localization of the Y determinant, i.e. Fucα(1→2)Galβ(1→4)[Fucα(1→3)]GlcNAcβ(1→·), in an N-glycan of a glycoprotein.

Immunologic Determination of Seminal Secretions by a Latex Microtiter Technique

The presence of human seminal plasma proteins in biological stains was demonstrated by an absorption test using anti-human seminal protein rabbit serum decreased degree of antibody activity was titrated on a microtiter plate by the agglutination of latex particles coated with human seminal plasma proteins. This method of test was sensitive and highly specific. The latex reagent for this test could be preserved in a refrigerator for over one year without the loss of reactivity.

Identification of species specific hemoglobin by isoelectric focusing

A method for the discrimination of human hemoglobin from animal sources is described. Hemoglobins from 34 species with and without treatment by p-chloromercuribenzoate (PCMB) were examined by thin-layer isoelectric focusing (IEF) in polyacrylamide gel (PAG) plates containing an Ampholine gradient of pH 3.5 - 9.5. PCMB treatment was effective in distinguishing human and animal hemoglobins with their characteristic IEF patterns. Values for the isoelectric points of the major and minor hemoglobins of each species are also described.