Conformational changes in human serum albumin around the neutral pH from circular dichroic measurements

Abstract

SummaryThe molar ellipticity of the warfarin-albumin complex at 310 nm increases with pH from 6 to 9. This pH dependence runs parallel with that of the molar ellipticity of the albumin alone at 292 nm. The change in molar ellipticity with pH occurs in a smaller pH interval after addition of the physiological concentration of calcium ions. These findings give support to the assumption that the binding site for warfarin on the albumin molecule is affected by the neutral-to-base transition in the protein.Several authors have reported a pH dependent conformational change to occur in serum albumin around the physiological pH [1--7]. Zurawski and Foster [ 2] showed evidence for the existence of two conformational states in bovine serum albumin, the N form mainly occurring at the neutral pH and the B form at higher pH. They call this conformational change, therefore, the neutral-to-base or the N-B transition. Leonard et al. [ 1 ] found that the specific rotation of bovine and human serum albumin at 313.2 nm decreases between pH 7 and 9. This was interpreted as a change of the protein structure to another form when the pH was increased. This basic form is also favoured with increasing ionic strength. Nikkel and Foster [7] and Zurawski and Foster [2] later showed similar effects of pH on the conformation of bovine serum albumin using NMR spectroscopy to follow the chemically modified albumin molecule. Katz and Klotz [8] and Harmsen et al. [3] showed for bovine serum albumin that this N-B transition was effected by calcium ions.

The role of the transition between neutral and basic forms of human serum albumin in the kinetics of the binding to warfarin

“…This result supports the hypothesis that the sensitivity of the value of Keq to the conformational state of albumin is dueto k2, which is the rate constant of the association reaction between worfarin and albumin leading to the stable complex. We found (data not shown) that Ca 2+ and C1-ions affect the pH dependence of k 2 in the same way as reported previously for the pH dependence of Keq [1,2]. This effect of Ca 2+ and C1-on the pH dependence of K~q was shown to be due to a change in the state of the N ~ B equilibrium by these ions.…”

“…This equation is valid, when the ligand molecule to be studied has no marked effect on the state of the N ~ B equilibrium on binding to the albumin molecule; warfarin fulfils this condition [1,2]. Since a can be derived from CD measurements, as described previously [2], the pH dependence of kob s can be calculated.…”

“…A pH-dependent conformational change in bovine and human serum albumin around physiological pH is often mentioned in the literature (see Wilting et al [1,2] for references). There is evidence for the existence of two conformational states, the N form occurring mainly below neutral pH and the B form at higher pH.…”

The role of albumin conformation in the binding of diazepam to human serum albumin

“…The first one was shown to be sensitive on the N-B transition [1,2]. The aim of this investigation is to demonstrate such a dependency for the diazepam binding site.…”

“…The molar ellipticity [0] increases with pH between pH 6 and 7.5, whereas [0] is constant out of this pH range. The pH range, over which [0] increases is at somewhat lower pH than the pH range, where the N-B transition is observed by means of CD using bound warfarin as a label [1,2]. The change in [0] with pH cannot be attributed to a change in physical chemical properties of the diazepam molecule, since the pK a of diazepam is 3.3 [20].…”