Abstract
Serum albumin possesses esterase and pseudo-esterase activities towards a number of endogenous and exogenous substrates, but the mechanism of interaction of various esters and other compounds with albumin is still unclear. In the present study, proton nuclear magnetic resonance (1H NMR) has been applied to the study of true esterase activity of albumin, using the example of bovine serum albumin (BSA) and p-nitrophenyl acetate (NPA). The site of BSA esterase activity was then determined using molecular modelling methods. According to the data obtained, the accumulation of acetate in the presence of BSA in the reaction mixture is much more intense as compared with the spontaneous hydrolysis of NPA, which indicates true esterase activity of albumin towards NPA. Similar results were obtained for p-nitophenyl propionate (NPP) as substrate. The rate of acetate and propionate release confirms the assumption that there is a site of true esterase activity in the albumin molecule, which is different from the site of the pseudo-esterase activity Sudlow II. The results of molecular modelling of BSA and NPA interaction make it possible to postulate that Sudlow site I is the site of true esterase activity of albumin.
Highlights
Albumin is the main protein in the blood of mammals, where its concentration is 500–700 μM
Cysteine was found to be the major molecule attached to the sulfydryl group of the cysteinyl residue of serum albumin [37], in our study, we studied the interaction of cysteinylated bovine serum albumin (BSA) with nitrophenyl acetate (NPA)
In this work, using the 1H NMR technique, we have demonstrated that serum albumin possesses true esterase activity
Summary
Albumin is the main protein in the blood of mammals, where its concentration is 500–700 μM. ABasAedccoonrdthinegtottoaliMtyD Simu tion of these data, we believe that it is Sudlow site I with catalytic Tyr149 (Tyr150 in HSA) that is the site of albumin true esterase activity. In the case of NPA hydrolysis by Sudlow site I of BSA, His241 can play the role of an oxyanion center, while Asp258 can attract the proton of the catalytic tyrosine. Earlier, based on modelling the interaction of albumin with OPs, we suggested that the catalytic “dyad” His-Tyr or Lys-Tyr, in which histidine or lysine acts as an acid residue, is required for the hydrolysis of some substrates with albumin [4] Both cases are described in the literature: the sedolisin proteases carry out catalysis using the Ser/Glu/Asp triad, while the Ser/His/His catalytic triad was validated by the crystal structure of the cytomegalovirus protease [44]. Quantitative analysis of the true esterase activity of albumin will help find a place for albumin in the enzyme nomenclature
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