Biomaterial Database

65Zn(II), 115mCd(II), 60Co(II), and mg(II) binding to alkaline phosphatase of Escherichia coli. Structural and functional effects. 1983

J E Coleman, and K Nakamura, and J F Chlebowski

Zn(II), Cd(II), Co(II) and Mg(II) binding to apoalkaline phosphatase of Escherichia coli and the relative stabilities of the resulting metalloenzyme complexes have been measured by equilibrium dialysis and metal exchange reactions using gamma-emitting isotopes of these metals. At millimolar concentrations of these metal ions the alkaline phosphatase dimer binds three pairs of metal ions (A, B, and C sites). One of these pairs dialyzes readily without detectable change in the structure or function of the enzyme (C site). Of the remaining two pairs, the binding affinity of both for Zn(II) and Cd(II) is increased by formation of the phosphoenzyme intermediates. Cd(II) is bound less tightly to both A and B sites than Zn(II), and at pH 6.5 Cd(II) is induced to bind to the B sites by formation of the phosphate complexes. Mg(II), 5-10 mM, competes successfully with the IIB metal ions for the second or lower affinity pair of binding sites (B sites), although Mg(II) is a relatively poor competitor on an equimolar basis, especially for Cd(II). Binding of metal ions to the apoenzyme appears to be a cooperative process involving conformational changes in the protein which are not readily reversible. The initial binding of a pair of Zn(II) or Cd(II) ions to the apoenzyme is characterized by equilibrium constants of 10(-5) to 10(-7) M for Zn(II) and 10(-4) to 10(-5) M for Cd(II). Following the cooperative binding of all three pairs of metal ions, however, re-establishment of equilibrium by dialysis indicates binding constants of less than 10(-8) M for Zn(II) and less than 10(-6) M for Cd(II) at the sites of greatest affinity (A sites). Binding of Mg(II) or Cd(II) to the B site, once the A site is occupied, increases the phosphorylation rate of the Cd(II) enzyme by 20-fold. In the presence of saturating concentrations of Mg(II) complete activity is restored to the apoenzyme by 2 Zn(II) ions. In the absence of Mg(II) as many as 6 Zn(II) ions may be required before complete restoration is achieved. Roles for the A and B site metal ions in the catalytic mechanism are discussed.

UI	MeSH Term	Description	Entries
D007700	Kinetics	The rate dynamics in chemical or physical systems.
D008274	Magnesium	A metallic element that has the atomic symbol Mg, atomic number 12, and atomic weight 24.31. It is important for the activity of many enzymes, especially those involved in OXIDATIVE PHOSPHORYLATION.
D011485	Protein Binding	The process in which substances, either endogenous or exogenous, bind to proteins, peptides, enzymes, protein precursors, or allied compounds. Specific protein-binding measures are often used as assays in diagnostic assessments.	Plasma Protein Binding Capacity,Binding, Protein
D002104	Cadmium	An element with atomic symbol Cd, atomic number 48, and atomic weight 112.41. It is a metal and ingestion will lead to CADMIUM POISONING.
D003035	Cobalt	A trace element that is a component of vitamin B12. It has the atomic symbol Co, atomic number 27, and atomic weight 58.93. It is used in nuclear weapons, alloys, and pigments. Deficiency in animals leads to anemia; its excess in humans can lead to erythrocytosis.	Cobalt-59,Cobalt 59
D004926	Escherichia coli	A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc.	Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D000469	Alkaline Phosphatase	An enzyme that catalyzes the conversion of an orthophosphoric monoester and water to an alcohol and orthophosphate. EC 3.1.3.1.
D001051	Apoenzymes	The protein components of enzyme complexes (HOLOENZYMES). An apoenzyme is the holoenzyme minus any cofactors (ENZYME COFACTORS) or prosthetic groups required for the enzymatic function.	Apoenzyme
D015032	Zinc	A metallic element of atomic number 30 and atomic weight 65.38. It is a necessary trace element in the diet, forming an essential part of many enzymes, and playing an important role in protein synthesis and in cell division. Zinc deficiency is associated with ANEMIA, short stature, HYPOGONADISM, impaired WOUND HEALING, and geophagia. It is known by the symbol Zn.