Catalytic role of the metal ion in the metallo-ß-lactamase GOB

TY - JOUR

T1 - Catalytic role of the metal ion in the metallo-ß-lactamase GOB

AU - Lise, María-Natalia

AU - Hemmingsen, Lars Bo Stegeager

AU - Vila, Alejandro J.

PY - 2010/2/12

Y1 - 2010/2/12

N2 - Metallo-β-lactamases (MβLs) stand as one of the main mechanisms of bacterial resistance toward carbapenems. The rational design of an inhibitor for MβLs has been limited by an incomplete knowledge of their catalytic mechanism and by the structural diversity of their active sites. Here we show that the MβL GOB from Elizabethkingia meningoseptica is active as a monometallic enzyme by using different divalent transition metal ions as surrogates of the native Zn(II) ion. Of the metal derivatives in which Zn(II) is replaced, Co(II) and Cd(II) give rise to the most active enzymes and are shown to occupy the same binding site as the native ion. However, Zn(II) is the only metal ion capable of stabilizing an anionic intermediate that accumulates during nitrocefin hydrolysis, in which the C-N bond has already been cleaved. This finding demonstrates that the catalytic role of the metal ion in GOB is to stabilize the formation of this intermediate prior to nitrogen protonation. This role may be general to all MβLs, whereas nucleophile activation by a Zn(II) ion is not a conserved mechanistic feature.

AB - Metallo-β-lactamases (MβLs) stand as one of the main mechanisms of bacterial resistance toward carbapenems. The rational design of an inhibitor for MβLs has been limited by an incomplete knowledge of their catalytic mechanism and by the structural diversity of their active sites. Here we show that the MβL GOB from Elizabethkingia meningoseptica is active as a monometallic enzyme by using different divalent transition metal ions as surrogates of the native Zn(II) ion. Of the metal derivatives in which Zn(II) is replaced, Co(II) and Cd(II) give rise to the most active enzymes and are shown to occupy the same binding site as the native ion. However, Zn(II) is the only metal ion capable of stabilizing an anionic intermediate that accumulates during nitrocefin hydrolysis, in which the C-N bond has already been cleaved. This finding demonstrates that the catalytic role of the metal ion in GOB is to stabilize the formation of this intermediate prior to nitrogen protonation. This role may be general to all MβLs, whereas nucleophile activation by a Zn(II) ion is not a conserved mechanistic feature.

U2 - 10.1074/jbc.M109.063743

DO - 10.1074/jbc.M109.063743

M3 - Journal article

C2 - 20007696

SN - 0021-9258

VL - 285

SP - 4570

EP - 4577

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

IS - 7

ER -