Structural Basis for Assembly of the MnIV/FeIII Cofactor in the Class Ic Ribonucleotide Reductase from Chlamydia trachomatis

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• 作者：Laura M. K. Dassama ; Carsten Krebs ; J. Martin Bollinger ; Jr. ; Amy C. Rosenzweig ; Amie K. Boal
• 刊名：Biochemistry
• 出版年：2013
• 出版时间：September 17, 2013
• 年：2013
• 卷：52
• 期：37
• 页码：6424-6436
• 全文大小：548K
• 年卷期：v.52,no.37(September 17, 2013)
• ISSN：1520-4995

文摘

The class Ic ribonucleotide reductase (RNR) from Chlamydia trachomatis (Ct) employs a Mn^IV/Fe^III cofactor in each monomer of its 尾₂ subunit to initiate nucleotide reduction. The cofactor forms by reaction of Mn^II/Fe^II-尾₂ with O₂. Previously, in vitro cofactor assembly from apo 尾₂ and divalent metal ions produced a mixture of two forms, with Mn at site 1 (Mn^IV/Fe^III) or site 2 (Fe^III/Mn^IV), of which the more active Mn^IV/Fe^III product predominates. Here we have addressed the basis for metal site selectivity by determining X-ray crystal structures of apo, Mn^II, and Mn^II/Fe^II complexes of Ct 尾₂. A structure obtained anaerobically with equimolar Mn^II, Fe^II, and apoprotein reveals exclusive incorporation of Mn^II at site 1 and Fe^II at site 2, in contrast to the more modest site selectivity achieved previously. Site specificity is controlled thermodynamically by the apoprotein structure, as only minor adjustments of ligands occur upon metal binding. Additional structures imply that, by itself, Mn^II binds in either site. Together, the structures are consistent with a model for in vitro cofactor assembly in which Fe^II specificity for site 2 drives assembly of the appropriately configured heterobimetallic center, provided that Fe^II is substoichiometric. This model suggests that use of a Mn^IV/Fe^III cofactor in vivo could be an adaptation to Fe^II limitation. A 1.8 脜 resolution model of the Mn^II/Fe^II-尾₂ complex reveals additional structural determinants for activation of the cofactor, including a proposed site for side-on (畏²) addition of O₂ to Fe^II and a short (3.2 脜) Mn^II鈥揊e^II interionic distance, promoting formation of the Mn^IV/Fe^IV activation intermediate.