Rad50/SMC proteins and ABC transporters: unifying concepts from high-resolution structures

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Abstract

ATP-binding cassette (ABC)-type ATPases are chemo-mechanical engines for diverse biological pathways. ABC ATPase domains act not only in ABC transporters but also in DNA mismatch, nucleotide excision and double-strand break repair enzymes, as well as in chromosome segregation. Atomic-resolution crystal structures suggest molecular mechanisms for ABC ATPases and reveal surprisingly significant mechanistic and architectural conservation. This emerging unified structural biochemistry provides general medical and biological insights into how ABC proteins function as chemo-mechanical devices. ATP binding by the signature and Q-loop motifs drives the conformations of substrate-specific domains to accomplish diverse functions in transmembrane transport and DNA repair.

Introduction

The ATP-binding cassette (ABC) is a ubiquitous, universally conserved ATPase domain that has historically been defined as the nucleotide-binding domain of an ABC transporter. ABC transporters comprise a large and diverse family of membrane-spanning proteins that specifically transport various substances, ranging from ions to proteins, across membranes 1., 2., 3.. With the availability of complete genomes and the refinement of bioinformatic tools, however, it has become apparent that ABC-type ATPase domains occur not only in ABC transporters but also in a variety of nontransporter proteins, such as structural maintenance of chromosome (SMC) proteins, the SMC-protein-related DNA double-strand break repair enzyme Rad50, the DNA mismatch repair enzyme MutS and the nucleotide excision repair enzyme UvrA [4].

ABC transporters and ABC family DNA repair enzymes are of exceptional biomedical interest, because members are implicated in human diseases and pathogen antibiotic resistance. Functional loss of the ABC transporter cystic fibrosis conductance regulator (CFTR) causes cystic fibrosis 5., 6. and the ABC transporter P-glycoprotein acts in the development of multidrug resistance by cancer cells 7., 8.. Mutations in the MSH (eukaryotic MutS homolog) DNA mismatch repair ABC ATPases are associated with hereditary nonpolyposis colon cancer [9]. Mutations in the DNA double-strand break repair multiprotein complex Rad50–Mre11–NBS1 result in the human cancer predisposition diseases Nijmegen breakage syndrome and ataxia telangiectasia variant 10., 11..

The recently discovered diversity in the occurrence of ABC-type ATPase domains is overshadowed by the extreme functional diversity between transport ABC ATPases and DNA repair/structural maintenance of chromosome ABC ATPases. In ABC transporters, the energy from ATP hydrolysis acts to transport specific substances across membranes [12]. In DNA mismatch repair, the MutS ABC ATPase utilizes ATP to recognize and bind mispaired DNA bases or DNA insertion loops 13., 14., 15.. This recognition is evidently a first step in postreplicative mismatch repair. In DNA double-strand break repair, the Rad50 ABC ATPase of the Mre11 complex 16., 17. uses ATP to bind and bridge DNA double-strand breaks, and to facilitate DNA end processing by the nuclease Mre11 18., 19., 20., 21., 22., 23.. In chromosome segregation and condensation by SMC ABC ATPases, ATP hydrolysis is essential, but its specific role in the gross conformational changes of sister chromatid cohesion and chromosome condensation is presently unclear [24].

Despite this remarkable functional diversity within and between transmembrane transport ABC ATPases and DNA repair ABC ATPases, it has recently become clear that all these ABC enzymes or multiprotein complexes share a similar architecture: two ABC ATPase domains with conserved ATP-binding motifs and signature motifs, as well as substrate- and function-specific domains. Moreover, high-resolution crystal structures of ABC enzymes from different families now allow us to unify the underlying fundamental ATP-driven conformational changes into a surprisingly conserved global conformational mechanism of ABC ATPases.

Section snippets

The conserved ABC ATPase domain fold and dimer assembly

High-resolution atomic structures of ABC ATPases are of considerable interest for revealing the basic mechanisms of this important ATPase enzyme family in human disease. Furthermore, there is interest in developing specific inhibitors aimed at targeting, for example, ABC ATPases that are involved in multidrug resistance 25., 26.. The first hallmark crystal structure of an ABC ATPase was the ABC ATPase domain of the bacterial histidine permease, HisP [27], which provided key information on the

The signature motif and quaternary conformational controls

One of the most conserved and yet puzzling motifs in ABC transporters has been the signature motif. The signature motif, also called the C-motif, consists of a conserved LSGGQ/E sequence. Significantly, this signature motif is as conserved as the ATP binding and hydrolyzing Walker A and B motifs; however, in the three-dimensional structure of ABC ATPase domains, the signature motif is remote from the ATP-binding region formed by the Walker A and B motifs [27]. However, mutations in the

Unified ABC architecture and allosteric control

How do these ATP-driven conformational changes within the ABC ATPase domain dimer drive the function of the complete, often multisubunit, ABC enzyme? Insights into this multicomponent allostery come from crystal structures of complete ABC enzymes (ButCD, MutS and MsbA) and from recent detailed structural and biochemical results for the Rad50–Mre11 complex 28.••, 33.••, 34.••, 35.••, 46.. A preserved feature of ABC enzymes is the tight association of the ABC ATPase domains with

SMC proteins and Rad50

Perhaps the most puzzling group of ABC enzymes is represented by the DNA repair protein Rad50 and the SMC family of proteins 16., 17., 24., 49., 50.. Rad50/SMC proteins are distinguished by an ABC ATPase domain with a more than 600-residue heptad repeat insertion, which forms a coiled coil. This domain organization leads to the striking three-dimensional architecture of Rad50 and SMC complexes, whereby the ABC ATPases form a globular head at the end of two large coiled-coil arms, which contain

Conclusions

Recent crystallographic and biochemical analyses of such diverse enzymes as ABC transporters, DNA repair enzymes and SMC ABC ATPases reveal basic mechanistic principles of great biological importance. Moreover, these results have uncovered an unexpected and remarkable conservation of ABC ATPase molecular mechanisms. First, ABC ATPases evidently function by a conserved engagement/disengagement cycle, in which ATP binding by the signature motif induces the tight dimerization of preoriented or

References and recommended reading

Papers of particular interest, published within the annual period of review, have been highlighted as:

  • of special interest

  • ••

    of outstanding interest

Acknowledgements

K-PH’s efforts are supported by the Deutsche Forschungsgemeinschaft (DFG) and the European Molecular Biology Organization (EMBO) young investigator program. The efforts of JAT and those of his laboratory are supported by the Office of Science, Department of Energy (DE-AC03-76SF00098) and the National Cancer Institute (P01 CA92584).

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