/sup 1/H NMR study of effects of synergistic anion and metal ion binding on pH titration of the histidinyl side-chain residues of the half-molecules of ovotransferrin

Journal Article · Tue Jun 02 00:00:00 EDT 1987 · Biochemistry; (United States)

OSTI ID:6124454

Woodworth, R C; Butcher, N D; Brown, S A; Brown-Mason, A

Separation of ovotransferrin into C-terminal (OTf/2C) and N-terminal (OTf/2N) half-molecules has made possible the resolution of all expected histidinyl C(2)H resonances by proton nuclear magnetic resonance at 250 MHz. The chemical shift of many of the resonances decreases with increasing pH, allowing construction of titration curves, whereas a few resonances fail to titrate. On formation of the Ga/sup III/OTf/2(C/sub 2/O/sub 4/) ternary complexes, two of the low-field C(2)H resonances in each half-molecule fail to titrate. This behavior implicates the imidazole groups giving rise to these resonances as ligands to the bound metal ion. A third C(2)H resonance in each half-molecule undergoes a marked reduction in pK'/sub a/ on formation of the ternary complex. The imidazole group displaying this resonance is implicated in a proton-relay scheme involved in binding the synergistic anion, oxalate, and a water of hydration on the bound metal ion. The titration curves for the various imidazole resonances have been fit to a four-parameter equation involving estimation of the pK'/sub a/, the limiting chemical shift values, and a Hill constant n. Hill constants of <1 can be rationalized by correcting the titration curve for the charge Z on the protein as a function of pH and the work function w. The titration curve for the imidazole group in OTf/2C involved in the proton-relay scheme shows a value for n > 1, which suggests positive cooperativity in the titration of this residue. The basis for this behavior cannot be rationalized at this time. /sup 13/C NMR studies of (zeta-/sup 13/C)Arg-OTf suggest the Arg side chains may not be intimately involved in formation of the ternary complex.

• Research Organization: Univ. of Vermont College of Medicine, Burlington
• OSTI ID: 6124454
• Journal Information: Biochemistry; (United States), Vol. 26:11
• Country of Publication: United States
• Language: English

Related Subjects

62 RADIOLOGY AND NUCLEAR MEDICINE
TRANSFERRIN
NUCLEAR MAGNETIC RESONANCE
CARBON 13
CHEMICAL SHIFT
HISTIDINE
IMIDAZOLES
IONS
NMR SPECTRA
PH VALUE
PROTONS
SYNERGISM
AMINO ACIDS
AZOLES
BARYONS
CARBON ISOTOPES
CARBOXYLIC ACIDS
CHARGED PARTICLES
ELEMENTARY PARTICLES
EVEN-ODD NUCLEI
FERMIONS
GLOBULINS
GLOBULINS-BETA
HADRONS
HETEROCYCLIC ACIDS
HETEROCYCLIC COMPOUNDS
ISOTOPES
LIGHT NUCLEI
MAGNETIC RESONANCE
METALLOPROTEINS
NUCLEI
NUCLEONS
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PROTEINS
RESONANCE
SPECTRA
STABLE ISOTOPES
550601* - Medicine- Unsealed Radionuclides in Diagnostics