AIM: The present study was undertaken to purify and partially characterize the 33.5-kilodalton (33.5 kDa) vesicular protein in human bile and to explore the possible molecular mechanisms of the initial crystal nucleat...AIM: The present study was undertaken to purify and partially characterize the 33.5-kilodalton (33.5 kDa) vesicular protein in human bile and to explore the possible molecular mechanisms of the initial crystal nucleation process.METHODS: The 33.5 kDa vesicular protein was isolated by ultracentrifugation and further purified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) under nonreducing conditions. The purified 33.5 kDa vesicular protein was subjected to N-terminal amino acid sequencing and amino acid analysis. Cholesterol crystallization activity was detected by cholesterol crystal growth assay. The sugar chain of the 33.5 kDa vesicular protein was analyzed by dot-immunobinding assay of lectin coupled to a peroxidase (HRP-DSA, HRP-ConA, HRP-WGA) and was deglycosylated using two different enzymatic approaches (N-deglycosylation and O-deglycosylation) to determine the molecular weight of the protein component, the type of linkage between polypeptide and carbohydrate components.RFSULTS: The 33.5 kDa vesicular protein with complicated glycan was an extensively glycosylated (37.3 %) monomer and these sugar chains strongly bound to DSA, but did not bind to ConA. Amino acid sequencing indicated that the protein was unique. The 33.5 kDa vesicular protein exhibited potent cholesterol crystallization promoting activity in vitro with derived crystal growth curve indices It, Ig, Ic presented as 0.57, 1.52, and 1.63 respectively. Both enzymatic proteolysis and N-deglycosylation of the protein removed all activity.CONCLUSION: These data suggest the 33.5 kDa vesicular protein may be responsible for the pathogenesis of cholesterol gallstone disease, and the sugar chains play an important role in pro-nucleating process.展开更多
There are many Zr particles in as-cast NiAl-33.5Cr-0.5Zr (at. pct) alloy, which usually exist at the edge of eutectic of beta -NiAl and cx-Cr. After air and furnace cooling solution treatments, far 1400 degreesC, 2 h ...There are many Zr particles in as-cast NiAl-33.5Cr-0.5Zr (at. pct) alloy, which usually exist at the edge of eutectic of beta -NiAl and cx-Cr. After air and furnace cooling solution treatments, far 1400 degreesC, 2 h and 1450 degreesC, 1 h, pure Zr phase remains in the furnace cooling (F.C.) state alloys and Ni2AlZr phase forms in the air cooling (A.C.) state alloys. During solution treatment at 1450 degreesC, bulk and 'fish bone' shape Zr-rich phases form respectively in F.C. and A.C. state alloys. A 'river' shape Ni2AlZr phase forms after 1450 C for 1h F.C. and 850 degreesC for 12 h, F.C.. The alloy has less pure Zr and Ni2AlZr phase after 1400 degreesC with both air and furnace cooling followed by 850 C and 950 C for 12 h, F.C. aging treatments, respectively. Additionally, there is a ternary eutectic of NiAlZr and a phase enriched Zr and Cr forms at the edge of the eutectic of beta -NiAl and alpha -Cr in the alloy treated at 1400 degreesC, 2 h, F.C. and 950 degreesC, 12 h, F.C.展开更多
基金the National Natural Science Foundation of China,No.30070737
文摘AIM: The present study was undertaken to purify and partially characterize the 33.5-kilodalton (33.5 kDa) vesicular protein in human bile and to explore the possible molecular mechanisms of the initial crystal nucleation process.METHODS: The 33.5 kDa vesicular protein was isolated by ultracentrifugation and further purified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) under nonreducing conditions. The purified 33.5 kDa vesicular protein was subjected to N-terminal amino acid sequencing and amino acid analysis. Cholesterol crystallization activity was detected by cholesterol crystal growth assay. The sugar chain of the 33.5 kDa vesicular protein was analyzed by dot-immunobinding assay of lectin coupled to a peroxidase (HRP-DSA, HRP-ConA, HRP-WGA) and was deglycosylated using two different enzymatic approaches (N-deglycosylation and O-deglycosylation) to determine the molecular weight of the protein component, the type of linkage between polypeptide and carbohydrate components.RFSULTS: The 33.5 kDa vesicular protein with complicated glycan was an extensively glycosylated (37.3 %) monomer and these sugar chains strongly bound to DSA, but did not bind to ConA. Amino acid sequencing indicated that the protein was unique. The 33.5 kDa vesicular protein exhibited potent cholesterol crystallization promoting activity in vitro with derived crystal growth curve indices It, Ig, Ic presented as 0.57, 1.52, and 1.63 respectively. Both enzymatic proteolysis and N-deglycosylation of the protein removed all activity.CONCLUSION: These data suggest the 33.5 kDa vesicular protein may be responsible for the pathogenesis of cholesterol gallstone disease, and the sugar chains play an important role in pro-nucleating process.
基金The work was supported by the National Advanced Materials Connittee of China(Grant No.970321016)the National Natural Science Foundation of Chind(No.59895152).
文摘There are many Zr particles in as-cast NiAl-33.5Cr-0.5Zr (at. pct) alloy, which usually exist at the edge of eutectic of beta -NiAl and cx-Cr. After air and furnace cooling solution treatments, far 1400 degreesC, 2 h and 1450 degreesC, 1 h, pure Zr phase remains in the furnace cooling (F.C.) state alloys and Ni2AlZr phase forms in the air cooling (A.C.) state alloys. During solution treatment at 1450 degreesC, bulk and 'fish bone' shape Zr-rich phases form respectively in F.C. and A.C. state alloys. A 'river' shape Ni2AlZr phase forms after 1450 C for 1h F.C. and 850 degreesC for 12 h, F.C.. The alloy has less pure Zr and Ni2AlZr phase after 1400 degreesC with both air and furnace cooling followed by 850 C and 950 C for 12 h, F.C. aging treatments, respectively. Additionally, there is a ternary eutectic of NiAlZr and a phase enriched Zr and Cr forms at the edge of the eutectic of beta -NiAl and alpha -Cr in the alloy treated at 1400 degreesC, 2 h, F.C. and 950 degreesC, 12 h, F.C.
