Exploring the range of protein flexibility, from a structural proteomics perspective

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Abstract

Changes in protein conformation play a vital role in biochemical processes, from biopolymer synthesis to membrane transport. Initial systematizations of protein flexibility, in a database framework, concentrated on the movement of domains and linkers. Movements were described in terms of simple sliding and hinging mechanisms of individual secondary structural elements. Recently, the accelerated pace and sophistication of methods for structural characterization of proteins has allowed high-resolution studies of increasingly complex assemblies and conformational changes. New data emphasize a breadth of possible structural mechanisms, particularly the ability to drastically alter protein architecture and the native flexibility of many structures.

Introduction

Annotations in the Database of Macromolecular Movements (http://molmovdb.org) 1., 2. currently include more than 240 distinct protein motions, the majority of which can be directly visualized from solved structures [3]. Domain motions of single subunits make up the largest subset, but an increasing number of molecular complexes, exhibiting large structural rearrangements have been solved. Initial attempts to classify motions used a convention of ‘shear’ versus ‘hinge’ movements [4] (based on the presence or absence of a maintained interface between moving parts) and typically focused on movements in single domains or large fragments. The repertoire of protein conformational changes has grown considerably, incorporating many cooperative movements of subunits and structural changes at a quaternary level, largely due to improvements in methods for structural characterization of large molecules. Efforts to computationally model the activity of macromolecular assemblages remain limited by time constraints, but recent studies have used simulation to investigate the global conformational changes of immense structures, such as the F1-ATPase [5], GroEL [6], and the 70S ribosome [7].

Here, we summarize several recent structural studies that illustrate the importance and diversity of protein motions, concentrating, primarily, on several groups of related protein structures or mechanisms. Although subtle conformational changes, down to the level of alternating sidechain rotamers, are often essential to protein function, our fundamental focus is on more global changes, involving significant movement of the protein backbone, and interactions between tertiary and quaternary elements. Furthermore, many of these changes might involve multiple distinct intermediate states or occur on time scales that are too large to permit conventional simulation. In particular, we have highlighted proteins that display considerable ‘plasticity’ or ‘fluidity’, in terms of changes in fold, interactions within the cell membrane, or movement in the native form.

An overview of several new motions, examined in the context of the database, is presented in Table 1 and Figure 1. With the exception of ATP sulfurylase, each structure listed has only a single chain that exhibits the described motion (although other subunits may be involved). Identical methods were used for gathering all of the presented statistics, but the nature of the structural changes varies widely, and most changes do not easily fall into one of the pre-existing categories; however, all of the structures shown have one or more flexible linker regions of multiple residues, from which much of the displacement is derived, and, although there are several cases of shearing helices, mobile interfaces are not usually maintained within a single chain.

Section snippets

T7 RNA polymerase

One of the most dramatic conformational changes that has been observed so far is seen in the elongation-phase structure of T7 RNA pol (Figure 1a). Although movement of some type is observed in RNA and DNA polymerases from a variety of organisms (for example, see [8] for a review of bacterial RNA polymerase structures), these typically involve flexible linkers between distinct rigid domains. The transition from initiation to elongation in the T7 polymerase requires refolding and massive

Membrane proteins

The improvement of techniques for structural characterization of membrane proteins has yielded several examples of structural changes in gating and transport, some involving considerable flexibility within the transmembrane region. An example of receptor functioning via conformational change was found in the structures of FecA [20], where ligand binding alters the conformation of extracellular loops, transmitting the signal to cytoplasmic proteins; but more complex motions are observed in

Ring complexes

Several known or suspected motions occur in complexes of identical subunits, arranged in a ring, whose motion is essentially cooperative. The best studied of these are GroEL, whose conformational cycle has been investigated by a host of biophysical techniques (including simulation), and aspartate transcarbamoylase. Recently, three new, completely unrelated structures of hexameric ATPases have been described, whose functionality depends on the conformation of the individual subunits.

Other structures

Several other structures that are not readily classified deserve mention here, particularly in the context of the inherently dynamic complexes described above. As in VirB11, evidence for multiple conformations is frequently found in single crystals, where two or more molecules in the asymmetric unit adopt different domain orientations. The two α subunits of the tetrameric Acetyl-CoA synthase (Figure 1g) are iron–sulfur binding proteins with three large domains, connected by hinges. In the

Conclusions

Theoretical studies of protein motion have traditionally focused on structures of single molecules, following a known transition (for example, domain closure in response to ligand binding [39]), or on detailed mechanistic and energetic analyses using simulation. Comparison of multiple structures is limited by available CPU power and by the diversity of tertiary arrangements; nevertheless, some trends might be seen by a proteomics approach. The degree of movement in many of the structures that

Supplementary material

Most of the structures discussed, for which 3D data are available, are listed online, at http://molmovdb.org/molmovdb/cocb. These listings include additional images and animations.

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

We thank Thomas Steitz and Whitney Yin for discussions of T7 polymerase function, and Michael Barnett, Duncan Milburn and Vadim Alexandrov for assistance with the database. We acknowledge support from the NIH (5P50 GM062413) and apologize to the many authors whose work could not be included due to space constraints.

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