INSIGHTS INTO THERMAL-RESISTANCE OF PROTEINS FROM THE INTRINSIC STABILITY OF THEIR ALPHA-HELICES

To investigate the role of alpha helices in protein thermostability, w e compared energy characteristics of alpha helices from thermophilic a nd mesophilic proteins belonging to four protein families of known thr ee-dimensional structure, for at least one member of each family. The changes in intrinsic free energy of alpha-helix formation were estimat ed using the statistical mechanical theory for describing helix/coil t ransitions in peptide helices [Munoz, V., Serrano, L. Nature Struc. Bi ol. 1:399-409, 1994; Munoz, V., Serrano, L. J. Mel. Biol. 245:275-296, 1995; Munoz, V., Serrano, L. J. Mel. Biol. 245:297-308, 1995]. Based on known sequences of mesophilic and thermophilic RecA proteins we fou nd that (1) a high stability of a helices is necessary but is not a su fficient condition for thermostability of RecA proteins, (2) the total helix stability, rather than that of individual helices, is the facto r determining protein thermostability, and (3) two facets of intraheli cal interactions, the intrinsic helical propensities of amino acids an d the side chain-side chain interactions, are the main contributors to protein thermostability. Similar analysis applied to families of L-la ctate dehydrogenases, seryl-tRNA synthetases, and aspartate amino tran sferases led us to conclude that an enhanced total stability of alpha helices is a general feature of many thermophilic proteins. The magnit ude of the observed decrease in intrinsic free energy on alpha-helix f ormation of several thermoresistant proteins was found to be sufficien t to explain the experimentally determined increase of their thermosta bility. Free energies of intrahelical interactions of different RecA p roteins calculated at three temperatures that are thought to be close to its normal environmental conditions were found to be approximately equal. This indicates that certain flexibility of RecA protein structu re is an essential factor for protein function. All RecA proteins anal yzed fell into three temperature-dependent classes of similar alpha-he lix stability (Delta G(int) = 45.0 +/- 2.0 kcal/mol). These classes we re consistent with the natural origin of the proteins. Based on the se quences of protein alpha helices with optimized arrangement of stabili zing interactions, a natural reserve of RecA protein thermoresistance was estimated to be sufficient for conformational stability of the pro tein at nearly 200 degrees C. (C) 1997 Wiley-Liss, Inc.