Tsang, W.Y., Ahmed, Naveed, Hinchliffe, Paul S., Wood, J. Matthew, Harding, Lindsay P., Laws, Andrew P. and Page, Michael I. (2005) Different transition-state structures for the reactions of β-lactams and analogous β-sultams with serine β-lactamases. Journal of the American Chemical Society, 127 (49). pp. 17556-17564. ISSN 1520-5126
Abstract

β-Sultams are the sulfonyl analogues of β-lactams, and N-acyl β-sultams are novel inactivators
of the class C β-lactamase of Enterobacter cloacaeP99. They sulfonylate the active site serine residue to
form a sulfonate ester which subsequently undergoes C-O bond fission and formation of a dehydroalanine
residue by elimination of the sulfonate anion as shown by electrospray ionization mass spectroscopy. The
analogous N-acyl β-lactams are substrates for β-lactamase and undergo enzyme-catalyzed hydrolysis
presumably by the normal acylation-deacylation process. The rates of acylation of the enzyme by the
β-lactams, measured by the second-order rate constant for hydrolysis, kcat/ Km, and those of sulfonylation
by the β-sultams, measured by the second-order rate constant for inactivation, ki
, both show a similar pH
dependence to that exhibited by the β-lactamase-catalyzed hydrolysis of β-lactam antibiotics. Electronwithdrawing
groups in the aryl residue of the leaving group of N-aroyl β-lactams increase the rate of alkaline
hydrolysis and give a Bronsted βlg of -0.55, indicative of a late transition state for rate-limiting formation
of the tetrahedral intermediate. Interestingly, the corresponding Bronsted βlg for the β-lactamase-catalyzed
hydrolysis of the same substrates is -0.06, indicative of an earlier transition state for the enzyme-catalyzed
reaction. By contrast, although the Bronsted βlg for the alkaline hydrolysis of N-aroyl β-sultams is -0.73,
similar to that for the β-lactams, that for the sulfonylation of β-lactamase by these compounds is -1.46,
compatible with significant amide anion expulsion/S-N fission in the transition state. In this case, the enzyme
reaction displays a later transition state compared with hydroxide-ion-catalyzed hydrolysis of the β-sultam.

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