Structural basis for assembly of the MnIV/FeIII cofactor in the class ic ribonucleotide reductase from chlamydia trachomatis

Laura M.K. Dassama, Carsten Krebs, Joseph M. Bollinger, Jr., Amy C. Rosenzweig, Amie Kathleen Boal

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

The class Ic ribonucleotide reductase (RNR) from Chlamydia trachomatis (Ct) employs a MnIV/FeIII cofactor in each monomer of its β2 subunit to initiate nucleotide reduction. The cofactor forms by reaction of MnII/FeII2 with O 2. Previously, in vitro cofactor assembly from apo β2 and divalent metal ions produced a mixture of two forms, with Mn at site 1 (MnIV/FeIII) or site 2 (FeIII/MnIV), of which the more active MnIV/FeIII product predominates. Here we have addressed the basis for metal site selectivity by determining X-ray crystal structures of apo, MnII, and MnII/Fe II complexes of Ct β2. A structure obtained anaerobically with equimolar MnII, FeII, and apoprotein reveals exclusive incorporation of MnII at site 1 and FeII 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, MnII binds in either site. Together, the structures are consistent with a model for in vitro cofactor assembly in which FeII specificity for site 2 drives assembly of the appropriately configured heterobimetallic center, provided that FeII is substoichiometric. This model suggests that use of a MnIV/Fe III cofactor in vivo could be an adaptation to FeII limitation. A 1.8 Å resolution model of the MnII/Fe II2 complex reveals additional structural determinants for activation of the cofactor, including a proposed site for side-on (η2) addition of O2 to FeII and a short (3.2 Å) MnII-FeII interionic distance, promoting formation of the MnIV/FeIV activation intermediate.

Original languageEnglish (US)
Pages (from-to)6424-6436
Number of pages13
JournalBiochemistry
Volume52
Issue number37
DOIs
StatePublished - Sep 17 2013

Fingerprint

Ribonucleotide Reductases
Chlamydia trachomatis
Apoproteins
Metals
Chemical activation
Metal ions
Nucleotides
Monomers
Crystal structure
X-Rays
Ions
Ligands
X rays
In Vitro Techniques

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

@article{c1b9f9d2cdfa43d082543c801461983e,
title = "Structural basis for assembly of the MnIV/FeIII cofactor in the class ic ribonucleotide reductase from chlamydia trachomatis",
abstract = "The class Ic ribonucleotide reductase (RNR) from Chlamydia trachomatis (Ct) employs a MnIV/FeIII cofactor in each monomer of its β2 subunit to initiate nucleotide reduction. The cofactor forms by reaction of MnII/FeII-β2 with O 2. Previously, in vitro cofactor assembly from apo β2 and divalent metal ions produced a mixture of two forms, with Mn at site 1 (MnIV/FeIII) or site 2 (FeIII/MnIV), of which the more active MnIV/FeIII product predominates. Here we have addressed the basis for metal site selectivity by determining X-ray crystal structures of apo, MnII, and MnII/Fe II complexes of Ct β2. A structure obtained anaerobically with equimolar MnII, FeII, and apoprotein reveals exclusive incorporation of MnII at site 1 and FeII 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, MnII binds in either site. Together, the structures are consistent with a model for in vitro cofactor assembly in which FeII specificity for site 2 drives assembly of the appropriately configured heterobimetallic center, provided that FeII is substoichiometric. This model suggests that use of a MnIV/Fe III cofactor in vivo could be an adaptation to FeII limitation. A 1.8 {\AA} resolution model of the MnII/Fe II-β2 complex reveals additional structural determinants for activation of the cofactor, including a proposed site for side-on (η2) addition of O2 to FeII and a short (3.2 {\AA}) MnII-FeII interionic distance, promoting formation of the MnIV/FeIV activation intermediate.",
author = "Dassama, {Laura M.K.} and Carsten Krebs and {Bollinger, Jr.}, {Joseph M.} and Rosenzweig, {Amy C.} and Boal, {Amie Kathleen}",
year = "2013",
month = "9",
day = "17",
doi = "10.1021/bi400819x",
language = "English (US)",
volume = "52",
pages = "6424--6436",
journal = "Biochemistry",
issn = "0006-2960",
publisher = "American Chemical Society",
number = "37",

}

Structural basis for assembly of the MnIV/FeIII cofactor in the class ic ribonucleotide reductase from chlamydia trachomatis. / Dassama, Laura M.K.; Krebs, Carsten; Bollinger, Jr., Joseph M.; Rosenzweig, Amy C.; Boal, Amie Kathleen.

In: Biochemistry, Vol. 52, No. 37, 17.09.2013, p. 6424-6436.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Structural basis for assembly of the MnIV/FeIII cofactor in the class ic ribonucleotide reductase from chlamydia trachomatis

AU - Dassama, Laura M.K.

AU - Krebs, Carsten

AU - Bollinger, Jr., Joseph M.

AU - Rosenzweig, Amy C.

AU - Boal, Amie Kathleen

PY - 2013/9/17

Y1 - 2013/9/17

N2 - The class Ic ribonucleotide reductase (RNR) from Chlamydia trachomatis (Ct) employs a MnIV/FeIII cofactor in each monomer of its β2 subunit to initiate nucleotide reduction. The cofactor forms by reaction of MnII/FeII-β2 with O 2. Previously, in vitro cofactor assembly from apo β2 and divalent metal ions produced a mixture of two forms, with Mn at site 1 (MnIV/FeIII) or site 2 (FeIII/MnIV), of which the more active MnIV/FeIII product predominates. Here we have addressed the basis for metal site selectivity by determining X-ray crystal structures of apo, MnII, and MnII/Fe II complexes of Ct β2. A structure obtained anaerobically with equimolar MnII, FeII, and apoprotein reveals exclusive incorporation of MnII at site 1 and FeII 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, MnII binds in either site. Together, the structures are consistent with a model for in vitro cofactor assembly in which FeII specificity for site 2 drives assembly of the appropriately configured heterobimetallic center, provided that FeII is substoichiometric. This model suggests that use of a MnIV/Fe III cofactor in vivo could be an adaptation to FeII limitation. A 1.8 Å resolution model of the MnII/Fe II-β2 complex reveals additional structural determinants for activation of the cofactor, including a proposed site for side-on (η2) addition of O2 to FeII and a short (3.2 Å) MnII-FeII interionic distance, promoting formation of the MnIV/FeIV activation intermediate.

AB - The class Ic ribonucleotide reductase (RNR) from Chlamydia trachomatis (Ct) employs a MnIV/FeIII cofactor in each monomer of its β2 subunit to initiate nucleotide reduction. The cofactor forms by reaction of MnII/FeII-β2 with O 2. Previously, in vitro cofactor assembly from apo β2 and divalent metal ions produced a mixture of two forms, with Mn at site 1 (MnIV/FeIII) or site 2 (FeIII/MnIV), of which the more active MnIV/FeIII product predominates. Here we have addressed the basis for metal site selectivity by determining X-ray crystal structures of apo, MnII, and MnII/Fe II complexes of Ct β2. A structure obtained anaerobically with equimolar MnII, FeII, and apoprotein reveals exclusive incorporation of MnII at site 1 and FeII 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, MnII binds in either site. Together, the structures are consistent with a model for in vitro cofactor assembly in which FeII specificity for site 2 drives assembly of the appropriately configured heterobimetallic center, provided that FeII is substoichiometric. This model suggests that use of a MnIV/Fe III cofactor in vivo could be an adaptation to FeII limitation. A 1.8 Å resolution model of the MnII/Fe II-β2 complex reveals additional structural determinants for activation of the cofactor, including a proposed site for side-on (η2) addition of O2 to FeII and a short (3.2 Å) MnII-FeII interionic distance, promoting formation of the MnIV/FeIV activation intermediate.

UR - http://www.scopus.com/inward/record.url?scp=84884256293&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84884256293&partnerID=8YFLogxK

U2 - 10.1021/bi400819x

DO - 10.1021/bi400819x

M3 - Article

C2 - 23924396

AN - SCOPUS:84884256293

VL - 52

SP - 6424

EP - 6436

JO - Biochemistry

JF - Biochemistry

SN - 0006-2960

IS - 37

ER -