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Despite their high sequence homology, rubredoxins fromDesulfovibrio gigasandD. desulfuricansare stabilized to very different extents by com-patible solutes such as diglycerol phosphate, the major osmolyte in the hyperthermophilic archaeonArchaeoglobus 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 fromD. desulfuricans | ềFEBS Journal Structural determinants of protein stabilization by solutes The importance of the hairpin loop in rubredoxins Tiago M. Pais1 Pedro Lamosa1 Wagner dos Santos1 Jean LeGall1 2 David L. Turner1 3 and Helena Santos1 1 Institute de Tecnologia Quimica e Biologica Universidade Nova de Lisboa Portugal 2 Department of Biochemistry University of Georgia Athens GA USA 3 Department of Chemistry University of Southampton UK Keywords compatible solutes hairpin structure NMR rubredoxin thermostability Correspondence H. Santos Instituto de Tecnologia Quimica e Biologica Universidade Nova de Lisboa Apartado 127 2780-156 Oeiras Portugal Fax 351 21 4428766 Tel 351 21 4469828 E-mail santos@itqb.unl.pt Received 22 June 2004 revised 7 December 2004 accepted 17 December 2004 doi 10.1111 j.1742-4658.2004.04534.x 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 Microbiol 66 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 A17 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 .
