Structural determinants of protein stabilization by solutes

Abstract

Despite their high sequence homology, rubredoxins from Desulfovibrio gigas and D. desulfuricans are stabilized to very different extents by compatible solutes such as diglycerol phosphate, the major osmolyte in the hyperthermophilic archaeon Archaeoglobus fulgidus[Lamosa P, Burke A, Peist R, Huber R, Liu M Y, Silva G, Rodrigues‐Pousada C, LeGall J, Maycock C and Santos H (2000) Appl Environ Microbiol66, 1974–1979]. The principal structural difference between these two proteins is the absence of the hairpin loop in the rubredoxin from D. desulfuricans. Therefore, mutants of D. gigas rubredoxin bearing deletions in the loop region were constructed to investigate the importance of this structural feature on protein intrinsic stability, as well as on its capacity to undergo stabilization by compatible solutes. The three‐dimensional structure of the mutant bearing the largest deletion, Δ17|29, was determined by 1H‐NMR, demonstrating that, despite the drastic deletion, the main structural features were preserved. The dependence of the NH chemical shifts on temperature and solute concentration (diglycerol phosphate or mannosylglycerate) provide evidence of subtle conformational changes induced by the solute. The kinetic stability (as assessed from the absorption decay at 494 nm) of six mutant rubredoxins was determined at 90 °C and the stabilizing effect exerted by both solutes was assessed. The extent of protection conferred by each solute was highly dependent on the specific mutant examined: while the half‐life for iron release in the wild‐type D. gigas rubredoxin increased threefold in the presence of 0.1 m diglycerol phosphate, mutant Δ23|29 was destabilized. This study provides evidence for solute‐induced compaction of the protein structure and occurrence of weak, specific interactions with the protein surface. The relevance of these findings to our understanding of the molecular basis for protein stabilization is discussed.

Relationship between Protein Stabilization and Protein Rigidification Induced by Mannosylglycerate

“…Even more surprisingly, the degree of stabilization rendered by a particular solute on a series of single mutant proteins varied significantly. 12 These results are difficult to explain with any of the models proposed for the stabilization of proteins by solutes. Putative specific interactions with the surface of the native protein might account for the observed effects, but their existence lacks clear evidence.…”

Organic compatible solutes of halotolerant and halophilic microorganisms

“…As another example, two archaeal rubredoxins have been shown to be stabilized to quite different extents by α-diglycerol phosphate [150]. Their structures are similar, except that one is missing a hairpin loop.…”

A unique glyceryl diglycoside identified in the thermophilic, radiation-resistant bacterium Rubrobacter xylanophilus

“…For instance, MGlyG at 0.25 M concentration increased the melting temperature of SNase by 4.5 °C more than the protection provided by MG, but if we consider malate dehydrogenase, both solutes display identical increases in the melting temperature of this enzyme. This apparent protein/solute pair specificity has been previously observed in several systems (Lamosa et al 2000;Faria et al 2008;Pais et al 2005).…”