期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2002
卷号：99
期号：23
页码：14752-14757
DOI：10.1073/pnas.232361199
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：The catalytic cycle of the ATP/Mg2+-dependent enzyme {beta}-lactam synthetase ({beta}-LS) from Streptomyces clavuligerus has been observed through a series of x-ray crystallographic snapshots. Chemistry is initiated by the ordered binding of ATP/Mg2+ and N2-(carboxyethyl)-L-arginine (CEA) to the apoenzyme. The apo and ATP/Mg2+ structures described here, along with the previously described CEA*,{beta}-methyleneadenosine 5'-triphosphate (CEA*AMP-CPP)/Mg2+ structure, illuminate changes in active site geometry that favor adenylation. In addition, an acyladenylate intermediate has been trapped. The substrate analog N2-(carboxymethyl)-L-arginine (CMA) was adenylated by ATP in the crystal and represents a close structural analog of the previously proposed CEA-adenylate intermediate. Finally, the structure of the ternary product complex deoxyguanidinoproclavaminic acid (DGPC)*AMP/PPi/Mg2+ has been determined. The CMA-AMP/PPi/Mg2+ and DGPC*AMP/PPi/Mg2+ structures reveal interactions in the active site that facilitate {beta}-lactam formation. All of the ATP-bound structures differ from the previously described CEA*AMP-CPP/Mg2+ structure in that two Mg2+ ions are found in the active sites. These Mg2+ ions play critical roles in both the adenylation and {beta}-lactamization reactions.
