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Titolo:
Human recombinant interferon-inducible protein-10: Intact disulfide bridges are not required for inhibition of hematopoietic progenitors and chemotaxis of T lymphocytes and monocytes
Autore:
Crow, M; Taub, DD; Cooper, S; Broxmeyer, HE; Sarris, AH;
Indirizzi:
Univ Texas, MD Anderson Canc Ctr, Dept Lymphoma Myeloma, Houston, TX 77030USA Univ Texas Houston TX USA 77030 pt Lymphoma Myeloma, Houston, TX 77030USA NCI, Frederick Canc Res & Dev Ctr, Frederick, MD 21702 USA NCI Frederick MD USA 21702 ck Canc Res & Dev Ctr, Frederick, MD 21702 USA Indiana Univ, Sch Med, Dept Med, Indianapolis, IN 46202 USA Indiana Univ Indianapolis IN USA 46202 pt Med, Indianapolis, IN 46202 USA Indiana Univ, Sch Med, Dept Immunol Microbiol, Indianapolis, IN 46202 USA Indiana Univ Indianapolis IN USA 46202 robiol, Indianapolis, IN 46202 USA Indiana Univ, Sch Med, Walther Oncol Ctr, Indianapolis, IN 46202 USA Indiana Univ Indianapolis IN USA 46202 ol Ctr, Indianapolis, IN 46202 USA
Titolo Testata:
JOURNAL OF HEMATOTHERAPY & STEM CELL RESEARCH
fascicolo: 1, volume: 10, anno: 2001,
pagine: 147 - 156
SICI:
1525-8165(200102)10:1<147:HRIPID>2.0.ZU;2-Y
Fonte:
ISI
Lingua:
ENG
Soggetto:
IN-VIVO; 3-DIMENSIONAL STRUCTURE; CHEMOKINE RECEPTORS; GEL ELECTROPHORESIS; SULFHYDRYL GROUPS; ESCHERICHIA-COLI; INTERLEUKIN-8; CELLS; IP-10; EXPRESSION;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Clinical Medicine
Life Sciences
Citazioni:
41
Recensione:
Indirizzi per estratti:
Indirizzo: Sarris, AH Univ Texas, MD Anderson Canc Ctr, Dept Lymphoma Myeloma, 1515 Holcombe Blvd, Houston, TX 77030 USA Univ Texas 1515 Holcombe Blvd Houston TX USA 77030 TX 77030 USA
Citazione:
M. Crow et al., "Human recombinant interferon-inducible protein-10: Intact disulfide bridges are not required for inhibition of hematopoietic progenitors and chemotaxis of T lymphocytes and monocytes", J HEMATH ST, 10(1), 2001, pp. 147-156

Abstract

Human recombinant interferon-inducible protein-10 (rIP-10), a C-X-C chemokine, inhibits proliferation of human hematopoietic progenitors responsive to co-stimulation by recombinant steel factor (rSLF), is chemotactic for human monocytes and T-lymphocytes, and promotes T-lymphocyte adhesion to endothelial cells. Because chemokines have four conserved cysteines forming two intramolecular disulfide bridges, we decided to investigate their contribution in the biological activity of rIP-10. Since amino acid residues 22-98 of the sequence predicted by the cDNA constitute the naturally occurring IP-10, they were cloned after an initiating methionine into expression vector pET-3d. Subsequently rIP-10 was purified by enzymatic cell lysis, solubilization of refractile bodies with guanidine hydrochloride, renaturation by dialysis against dilute acetic acid, and sequential ion-exchange and reverse-phase high-performance liquid chromatography. Purified rIP-10 was reduced with 20 mM dithiothreitol, and chemically modified with 100 mM iodoacetamide(IAA), or S-methyl-methanethiosulfonate (MMTS), or N-methylmaleimide (NMM). Radiolabeling experiments demonstrated that 95% of the rIP-10 thiols weremodified, and this was confirmed with SDS-PAGE. The biological activity ofmodified rIP-10 was determined in vitro by inhibition of rSLF-responsive human bone marrow hematopoietic progenitor proliferation and by chemotaxis assays using human T-lymphocytes and monocytes. In both assay systems, the biological activity was evident at rIP-10 concentrations of 20-100 ng/ml. The activity was preserved after modification of rIP-10 by IAA or MMTS, but was abolished after modification by NMM. We conclude that disulfide bridges are not essential for the biological activity of rIP-10.

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Documento generato il 01/04/20 alle ore 20:51:56