Formation of the metal-thiolate clusters of rat liver metallothionein

Journal Article · Thu Mar 01 00:00:00 EST 1984 · Environ. Health Perspect.; (United States)

DOI:https://doi.org/10.1289/ehp.8454129· OSTI ID:5930778

Winge, D R; Nielson, K B

The isoforms of rat liver apo-metallothionein (MT) were reconstituted in vitro with Cd and Zn ions to study the order of binding of the seven metal sites. Reconstitution with seven Cd ions resulted in a metalloprotein similar to induced Cd, Zn-MT by the criteria of electrophoretic mobility, insensitivity to proteolysis by subtilisin and the pH-dependent release of Cd. Proteolytic digestion of MT reconstituted with sub-optimal quantities of Cd followed by separation of Cd-containing polypeptide fragments by electrophoresis and chromatography revealed metal ion binding initially occurs in cluster A. Upon saturation of the four sites in cluster A, binding occurs in the three metal center, cluster B. Samples reconstituted with one to four Cd or Zn ions per protein molecule, followed by digestion with subtilisin, yielded increasing amounts of a proteolytically stable polypeptide fragment identical with the ..cap alpha.. fragment domain encompassing the four metal center. Samples renatured with five to seven Cd ions per MT molecule showed decreasing quantities of ..cap alpha.. fragment and increasing amounts of native-like MT. The binding process in each domain is cooperative. Reconstitution of apo-MT with two Cd ions followed by proteolysis yields a 50% recovery of saturated Cd/sub 4/-..cap alpha.. cluster. Likewise, when Cd/sub 5/-renatured MT was digested with subtilisin, 30% of the molecules were identified as Cd/sub 7/-MT with the remainder as Cd/sub 4/-..cap alpha.. fragment. 11 references, 1 table, 3 figures.

• Research Organization: Univ. of Utah Medical Center, Salt Lake City
• OSTI ID: 5930778
• Journal Information: Environ. Health Perspect.; (United States), Vol. 54
• Country of Publication: United States
• Language: English

Related Subjects

63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.
59 BASIC BIOLOGICAL SCIENCES
METALLOTHIONEIN
BIOCHEMICAL REACTION KINETICS
STRUCTURAL CHEMICAL ANALYSIS
BIOLOGICAL PATHWAYS
CADMIUM
IN VITRO
LIVER
RATS
ZINC
ANIMALS
BODY
DIGESTIVE SYSTEM
ELEMENTS
GLANDS
KINETICS
MAMMALS
METALLOPROTEINS
METALS
ORGANIC COMPOUNDS
ORGANS
PROTEINS
REACTION KINETICS
RODENTS
VERTEBRATES
560305* - Chemicals Metabolism & Toxicology- Vertebrates- (-1987)
550200 - Biochemistry