1 | INTRODUCTION
Every two years since 1994, CASP (Critical Assessment of Structure Prediction) has conducted a community experiment to determine the state of the art in computing protein structure from amino acid sequence. Participants are provided with sequence information for non-public structures and invited to submit computed 3D structures which are then assessed by comparison with experimental data using established metrics. The 2020 (CASP14) experiment (1) saw the problem essentially solved for most single protein structures (2) using a new deep learning method, AlphaFold2 (AF2) (3). Many computed structures were competitive in accuracy with the corresponding experimental ones. To build on these results, the scope of CASP15 (2022) was expanded to include several new areas where the deep learning methods hold promise for further progress (4). One area of high significance and challenge is the modeling of multiple conformational states of macromolecules and macromolecular complexes. While a single conformation might be sufficient to understand function, in other situations, the ability of macromolecules to adopt multiple conformational states provides the basis for functional properties.
Multiple conformations can arise in a variety of situations. One class are those that occur under the same environmental conditions and these fall into two main groups: ordered or intrinsically disordered proteins /domains /regions. Disorder ensembles present unique computational challenges and are covered by one of CASP’s sister organizations, CAID (Critical Assessment of Intrinsic Disorder). The results of the most recent CAID round are also reported in this CASP special issue (5). Ordered multi-conformational structures were represented in CASP15 by low population states of three kinases (CASP targets T1195-T1197), investigated by NMR, and a long-lived folding intermediate of a structured RNA molecule (target R1138).
The second and more varied class of alternative conformations are those exerted under different conditions. Different conformational states may be induced in the presence of different small ligands, both organic and inorganic, or by binding to other macromolecules. Ligand binding effects on conformation are often allosteric in that they trigger conformational changes affecting function. For this class, CASP15 includes an ABC transporter target with four different ligands bound, inducing three primary conformational states (T1158 series) and an RNA target observed in two states, one with four protein molecules bound (R1190) and the other with six (R1189). Changes in the primary amino acid sequence, i.e. mutations, may also induce conformational changes, and there is one such case (T1109/10) where a single mutation induces a domain swap. Conformation may also change depending on functional state, and targets T1170, H1171, and H1172 are for different states of a Holliday junction complex. Solvent changes may also induce conformational variations and there is a pair of closely related targets (T1160/61) with conformations observed under different crystallization conditions.
Thus, although small in number, the CASP15 ensemble targets have provided a diverse set of multiple-conformational-state targets, and a very interesting pilot experiment in this area.