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121.
122.
Cutting edge: SR-PSOX/CXC chemokine ligand 16 mediates bacterial phagocytosis by APCs through its chemokine domain 总被引:11,自引:0,他引:11
Shimaoka T Nakayama T Kume N Takahashi S Yamaguchi J Minami M Hayashida K Kita T Ohsumi J Yoshie O Yonehara S 《Journal of immunology (Baltimore, Md. : 1950)》2003,171(4):1647-1651
SR-PSOX and CXC chemokine ligand (CXCL)16, which were originally identified as a scavenger receptor and a transmembrane-type chemokine, respectively, are indicated to be identical. In this study, we demonstrate that membrane-bound SR-PSOX/CXCL16 mediates adhesion and phagocytosis of both Gram-negative and Gram-positive bacteria. Importantly, our prepared anti-SR-PSOX mAb, which suppressed chemotactic activity of SR-PSOX, significantly inhibited bacterial phagocytosis by human APCs including dendritic cells. Various scavenger receptor ligands inhibited the bacterial phagocytosis of SR-PSOX. In addition, the recognition specificity for bacteria was determined by only the chemokine domain of SR-PSOX/CXCL16. Thus, SR-PSOX/CXCL16 may play an important role in facilitating uptake of various pathogens and chemotaxis of T and NKT cells by APCs through its chemokine domain. 相似文献
123.
Uyama T Kitagawa H Tanaka J Tamura J Ogawa T Sugahara K 《The Journal of biological chemistry》2003,278(5):3072-3078
We identified a novel human chondroitin N-acetylgalactosaminyltransferase, designated chondroitin GalNAcT-2 after a BLAST analysis of the GenBank(TM) data base using the sequence of a previously described human chondroitin N-acetylgalactosaminyltransferase (chondroitin GalNAcT-1) as a probe. The new cDNA sequence contained an open reading frame encoding a protein of 542 amino acids with a type II transmembrane protein topology. The amino acid sequence displayed 60% identity to that of human chondroitin GalNAcT-1. Like chondroitin GalNAcT-1, the expression of a soluble form of the protein in COS-1 cells produced an active enzyme, which not only transferred beta1,4-N-acetylgalactosamine (GalNAc) from UDP-[(3)H]GalNAc to a polymer chondroitin representing growing chondroitin chains (beta-GalNAc transferase II activity) but also to GlcUA beta 1-3Gal beta 1-O-C(2)H(4)NHCbz, a synthetic substrate for beta-GalNAc transferase I that transfers the first GalNAc to the core tetrasaccharide in the protein-linkage region of chondroitin sulfate. In contrast, the tetrasaccharide serine (GlcUA beta 1-3Gal beta 1-3Gal beta 1-4Xyl beta 1-O-Ser) derived from the linkage region, which is an inert acceptor substrate for chondroitin GalNAcT-1, served as an acceptor substrate. The coding region of this enzyme was divided into seven discrete exons, which is similar to the genomic organization of the chondroitin GalNAcT-1 gene, and was localized to chromosome 10q11.22. Northern blot analysis revealed that the chondroitin GalNAcT-2 gene exhibited a ubiquitous but differing expression in human tissues, and the expression pattern differed from that of chondroitin GalNAcT-1. Thus, we demonstrated redundancy in the chondroitin GalNAc transferases involved in the biosynthetic initiation and elongation of chondroitin sulfate, which is important for understanding the biosynthetic mechanisms leading to the selective chain assembly of chondroitin/dermatan sulfate on the linkage region tetrasaccharide common to various proteoglycans containing chondroitin/dermatan sulfate and heparin/heparan sulfate chains. 相似文献
124.
Koshizuka Y Yamada T Hoshi K Ogasawara T Chung UI Kawano H Nakamura Y Nakamura K Ikegawa S Kawaguchi H 《The Journal of biological chemistry》2003,278(48):48259-48266
This study attempts to characterize cystatin 10 (Cst10), which we recently identified as a novel protein implicated in endochondral ossification. Expression of Cst10 was specific to cartilage, localized in the cytosol of prehypertrophic and hypertrophic chondrocytes of the mouse growth plate. In the mouse chondrogenic cell line ATDC5, Cst10 expression preceded type X collagen expression and increased in synchrony with maturation. When we compared ATDC5 cells transfected with Cst10 cDNA with cells transfected with a mock vector, hypertrophic maturation and mineralization of chondrocytes were promoted by Cst10 gene overexpression in that type X collagen expression was observed earlier, and alizarin red staining was stronger. On the other hand, type II collagen expression and Alcian blue staining, both of which are markers of the early stage of chondrocyte differentiation, were similar in both cells. Overexpression of the Cst10 gene also caused fragmentation of nuclei, the appearance of annexin V, a change in the mitochondrial membrane potential, and activation of caspases. These results strongly suggest that Cst10 may play an important role in the last steps of the chondrocyte differentiation pathway as an inducer of maturation, followed by apoptosis of chondrocytes. 相似文献
125.
Yoshimura T Sagami I Sasakura Y Shimizu T 《The Journal of biological chemistry》2003,278(52):53105-53111
The heme-regulated phosphodiesterase (PDE) from Escherichia coli (Ec DOS) is a tetrameric protein composed of an N-terminal sensor domain (amino acids 1-201) containing two PAS domains (PAS-A, amino acids 21-84, and PAS-B, amino acids 144-201) and a C-terminal catalytic domain (amino acids 336-799). Heme is bound to the PAS-A domain, and the redox state of the heme iron regulates PDE activity. In our experiments, a H77A mutation and deletion of the PAS-B domain resulted in the loss of heme binding affinity to PAS-A. However, both mutant proteins were still tetrameric and more active than the full-length wild-type enzyme (140% activity compared with full-length wild type), suggesting that heme binding is not essential for catalysis. An N-terminal truncated mutant (DeltaN147, amino acids 148-807) containing no PAS-A domain or heme displayed 160% activity compared with full-length wild-type protein, confirming that the heme-bound PAS-A domain is not required for catalytic activity. An analysis of C-terminal truncated mutants led to mapping of the regions responsible for tetramer formation and revealed PDE activity in tetrameric proteins only. Mutations at a putative metal-ion binding site (His-590, His-594) totally abolished PDE activity, suggesting that binding of Mg2+ to the site is essential for catalysis. Interestingly, the addition of the isolated PAS-A domain in the Fe2+ form to the full-length wild-type protein markedly enhanced PDE activity (>5-fold). This activation is probably because of structural changes in the catalytic site as a result of interactions between the isolated PAS-A domain and that of the holoenzyme. 相似文献
126.
127.
Niimi S Oshizawa T Yamaguchi T Harashima M Seki T Ariga T Kawanishi T Hayakawa T 《Biochemical and biophysical research communications》2003,300(3):770-774
Small hepatocytes are cells that express characteristic phenotypes such as a high growth potential and differentiation capacity. In order to identify rat-small-hepatocyte specific proteins, we separated the cellular proteins of isolated small and parenchymal hepatocytes by 2D polyacrylamide gel electrophoresis. Comparison of their profiles revealed a protein with a molecular mass of 37 kDa in the small hepatocytes that was not present in the parenchymal hepatocytes. Proteolytic peptide mass fingerprinting was used to identify the protein and it was found to be annexin III. The validity of the identification was confirmed by Western blot analysis with anti-annexin III antibody. 相似文献
128.
The versatile plant acyltransferase (VPAT) family is a recently identified protein family consisting of acyltransferases involved in secondary metabolism in plants along with numerous homologues with as yet unidentified biochemical functions. Malonyl-CoA:anthocyanin 5-O-glucoside-6' "-O-malonyltransferase of Salvia splendens flowers (Ss5MaT1) is a member of this family that catalyzes the regiospecific transfer of the malonyl group from malonyl-CoA to the 6' "-hydroxyl group of the 5-glycosyl moiety of anthocyanins. To elucidate the mechanism and functional amino acid residues of VPAT family enzymes, steady-state kinetic analyses and site-directed mutagenesis of Ss5MaT1 guided by sequence comparison studies were carried out. On the basis of the results of product and dead-end inhibition studies as well as sequence comparison studies, the kinetic mechanism of Ss5MaT1 could be most consistently described in terms of a ternary complex mechanism in which both substrates and the enzyme form a complex before catalysis can occur, as in the case of chloramphenicol O-acetyltransferase (CAT) and histone acetyltransferase (HAT). Eight polar or ionizable amino acid residues that are invariant among 12 VPAT family enzymes were replaced by alanine, and the mutant enzymes were kinetically characterized. A significant diminution of the k(cat) value was observed with the substitution of His167 (relative k(cat), 0.02%) and Asp390 (<0.01%), strongly suggesting that His167 and Asp390 are very important for catalytic activity. The log k(cat) versus pH plots of the Ss5MaT1-catalyzed malonyl transfer suggested that a deprotonated active site group of pK(a) = 7.0 +/- 0.1 may be involved in the catalytic steps of the "substrate to product" conversion in the ternary enzyme-substrate complex. Taking these lines of evidence together with the suggested similarity of the kinetic and catalytic mechanisms of Ss5MaT1 to those of CAT and HAT, the following Ss5MaT1 mechanism based on general acid/base catalysis was proposed: in the ternary complex, a general base deprotonates the 6' "-hydroxyl group of the anthocyanin substrate, thereby promoting a nucleophilic attack on the carbonyl of the thioester of malonyl-CoA; His167 and Asp390 appear to be involved in the general acid/base mechanism of Ss5MaT1. 相似文献
129.
Human umbilical vein endothelium-derived cells retain potential to differentiate into smooth muscle-like cells 总被引:8,自引:0,他引:8
Mouse embryonic stem-derived cells were recently shown to differentiate into endothelial and smooth muscle cells. In the present study, we investigated whether human umbilical vein endothelium-derived cells retain the potential to differentiate into smooth muscle cells. Examination of biochemical markers, including basic calponin, SM22alpha, prostaglandin E synthase, von Willebrand factor, and PECAM-1, as well as cell contractility, showed that whereas endothelium-derived cells cultured with fibroblast growth factor can be characterized as endothelial cells, when deprived of fibroblast growth factor, a significant fraction differentiates into smooth muscle-like cells. Reapplication of fibroblast growth factor reversed this differentiation. Activin A was up-regulated in fibroblast growth factor-deprived, endothelium-derived cells; moreover, the inhibitory effects of exogenous follistatin and overexpressed Smad7 on smooth muscle-like differentiation confirmed that the differentiation was driven by activin A signaling. These findings indicate that when deprived of fibroblast growth factor, human umbilical vein endothelium-derived cells are capable of differentiating into smooth muscle-like cells through activin A-induced, Smad-dependent signaling, and that maintenance of the endothelial cell phenotype and differentiation into smooth muscle-like cells are reciprocally controlled by fibroblast growth factor-1 and activin A. 相似文献
130.
The putative Rhodococcus rrn promoter region was cloned from the benzothiophene desulfurizing Rhodococcus sp. strain T09, and the dibenzothiophene desulfurizing gene, dsz, was expressed under the control of the putative rrn promoter in the strain T09 using a Rhodococcus–E.coli shuttle vector. Strain T09 harboring the expression vector, pNT, could desulfurize dibenzothiophene in the presence of inorganic
sulfate, methionine, or cysteine, while the Dsz phenotype was completely repressed in recombinant cells carrying the gene
under the control of the native dsz promoter under the same conditions. Among the sulfur sources examined, no intermediates were detected and only the final
desulfurized product, 2-hydroxy-biphenyl, was produced using ammonium sulfate as the sulfur source.
Received: 4 December 2001 / Accepted: 7 January 2002 相似文献