We report on the existence of two kinds of matrix metalloproteinases (MMPs), MMP-2 and MMP-9, in human seminal plasma. Partial purification of the proteinases was achieved by two steps, consisting of chromatography on a gel-filtration column and then on a gelatin affinity column. Proteinase activities in the chromatography extracts were shown to hydrolyse a fluorescent substrate specific to MMPs (Dnp-Pro-Leu-Gly-Leu-Trp-Ala-D-Arg-NH2). The proteinases were detected using gelatin-zymography, but were not detected using casein-zymography, and were also inhibited by EDTA, EGTA and o-phenanthroline. Molecular weights of the proteinases were determined by SDS-PAGE, gelatin-zymography and Western blot to be approximately 92, 84, 72, 67, 52 and 45 kDa. Gelatin-zymography showed three major bands of activity at 72, 67 and 52 kDa and minor bands at 92, 84 and 45 kDa. Apart from the two smallest bands, these proteinases were all recognized by the polyclonal antibodies for MMP-2 or MMP-9. These results indicate that two kinds of pro-form and active-form matrix metalloproteinases, MMP-2 and MMP-9, and their degradation products, are present in human seminal plasma.

The thermal structure of the Sambagawa belt, which is reflected by minerals produced during the highest temperature of metamorphism, generally is characterized by the occurrence of the highest grade schists in the middle of the structural pile. The origin of this structure has been analysed in the upper horizon of the Sambagawa schist sequence in Central Shikoku as an example for the structure of the whole belt. The upper horizon of the Sambagawa schist sequence in Central Shikoku consists of three nappes, the Saruta nappe 11, the Saruta nappe I and the Fuyunose nappe in descending order of structural level. The Saruta nappe Il shows a downward increase of metamorphic grade from the garnet zone, through the albite-biotite zone, to the oligoclase-biotite zone. The Saruta nappe I consists mainly of rocks in the albite-biotite zone and partly in the oligoclase-biotite zone, but the direction of the increase of metamorphic grade is not clear. The Fuyunose nappe shows an upward increase in metamorphic grade which changes from the glaucophane zone to the barroisite zone. It is concluded that the Saruta nappe I1 and Saruta nappe I were overturned and then mechanically coupled with the Fuyunose nappe. The Sambagawa metamorphic field, which is of the highest temperature phase of metamorphism, appears to have had an inverted thermal gradient and a thermal structure comparable with that expected in the deeper parts of a subduction complex.

No abstract

The endoplasmic-reticulum-associated degradation of misfolded (glyco)proteins ensures that only functional, correctly folded proteins exit from the endoplasmic reticulum and that misfolded ones are degraded by the ubiquitin-proteasome system. During the degradation of misfolded glycoproteins, they are deglycosylated by the PNGase (peptide:N-glycanase). The free oligosaccharides released by PNGase are known to be further catabolized by a cytosolic alpha-mannosidase, although the gene encoding this enzyme has not been identified unequivocally. The findings in the present study demonstrate that an alpha-mannosidase, Man2C1, is involved in the processing of free oligosaccharides that are formed in the cytosol. When the human Man2C1 orthologue was expressed in HEK-293 cells, most of the enzyme was localized in the cytosol. Its activity was enhanced by Co2+, typical of other known cytosolic alpha-mannosidases so far characterized from animal cells. The down-regulation of Man2C1 activity by a small interfering RNA drastically changed the amount and structure of oligosaccharides accumulating in the cytosol, demonstrating that Man2C1 indeed is involved in free oligosaccharide processing in the cytosol. The oligosaccharide processing in the cytosol by PNGase, endo-beta-N-acetylglucosaminidase and alpha-mannosidase may represent the common 'non-lysosomal' catabolic pathway for N-glycans in animal cells, although the molecular mechanism as well as the functional importance of such processes remains to be determined.