期刊
JOURNAL OF INORGANIC BIOCHEMISTRY
卷 102, 期 3, 页码 489-499出版社
ELSEVIER SCIENCE INC
DOI: 10.1016/j.jinorgbio.2007.10.030
关键词
zinc; zinc trafficking; metallothionein; zinc-proteome; glutathione; transcription factor Sp1
资金
- NIEHS NIH HHS [ES-04026, P30 ES004184-21, P30 ES004184, ES-04184, R01 ES004026-21, R01 ES004026] Funding Source: Medline
- NIGMS NIH HHS [R01 GM085114] Funding Source: Medline
Many cell types contain metal-ion unsaturated metallothionein (MT). Considering the Zn2+ binding affinity of metallothionein, the existence of this species in the intracellular environment constitutes a substantial thermodynamic sink. Indeed, the mM concentration of glutathione may be thought of in the same way. In order to understand how apo-MT and the rest of the Zn-proteome manage to coexist, experiments examined the in vitro reactivity of Zn-proteome with apo-MT, glutathione (GSH), and a series of common Zn2+ chelating agents including N,N,N',N'-(2-pyridylethyl)ethylenediammine (TPEN), EDTA, and [(2,2'-oxyproplylene-dinitrilo]tetraacetic acid (EGTA). Less than 10% of Zn-proteome from U87 mg cells reacted with apo-MT or GSH. In contrast, each of the synthetic chelators was 2-3 times more reactive. TPEN, a cell permeant reagent, also reacted rapidly with both Zn-proteome and Zn-MT in LLC-PKI cells. Taking a specific zinc finger protein for further study, apo-MT, GSH, and TPEN inhibited the binding of Zn-3-SP1 with its cognate DNA site (GC-1) in the sodium-glucose co-transporter promoter of mouse kidney. In contrast, preformation of Zn-3-SP1-(GC-1) prevented reaction with apo-MT and GSH; TPEN remained active but at a higher concentration. Whereas, Zn-3-SP1 is active in cells containing apo-MT and GSH, exposure of LLC-PK1 cells to TPEN for 24 h largely inactivated its DNA binding activity. The results help to rationalize the steady state presence of cellular apo-MT in the midst of the many, diverse members of the Zn-proteome. They also show that TPEN is a robust intracellular chelator of proteomic Zn2+ (C) 2007 Elsevier Inc. All rights reserved.
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