caa a22 4500 445818557 CHVBK 20180317145236.0 cr unu---uuuuu 170323e20110501xx s 000 0 eng 10.1007/s00792-011-0363-6 doi (NATIONALLICENCE)springer-10.1007/s00792-011-0363-6 Contribution of inter-subunit interactions to the thermostability of Pyrococcus furiosus citrate synthase [Elektronische Daten] [Vicky Moore, Ami Kanu, Olwyn Byron, Gordon Campbell, Michael Danson, David Hough, Susan Crennell] Using citrate synthase from the hyperthermophile Pyrococcus furiosus (PfCS) as our test molecule, we show through guanidine hydrochloride-induced unfolding that the dimer separates into folded, but inactive, monomers before individual subunit unfolding takes place. Given that forces across the dimer interface are vital for thermostability, a robust computational method was derived that uses the University of Houston Brownian Dynamics (UHBD) program to calculate both the hydrophobic and electrostatic contribution to the dimerisation energy at 100°C. The results from computational and experimental determination of the lowered stability of interface mutants were correlated, being both of the same order of magnitude and placing the mutant proteins in the same order of stability. This computational method, optimised for hyperthermophilic molecules and tested in the laboratory, after further testing on other examples, could be of widespread use in the prediction of thermostabilising mutations in other oligomeric proteins for which dissociation is the first step in unfolding. Springer, 2011 Analytical ultracentrifugation nationallicence Fluorescence nationallicence UHBD nationallicence Dimerisation energy nationallicence Oligomeric stability nationallicence Citrate synthase nationallicence Archaeon nationallicence CS : Citrate synthase nationallicence DTNB : 5,5′-dithiobis(2-nitrobenzoic acid) nationallicence EPPS : 4-(2-Hydroxyethyl)-1-piperazinepropanesulfonic acid nationallicence GdnHCl : Guanidine hydrochloride nationallicence Pf CS : Pyrococcus furiosus citrate synthase nationallicence $$ s_{20,w}^{0} $$ : Sedimentation coefficient nationallicence SASA : Solvent-accessible surface area nationallicence UHBD : University of Houston Brownian Dynamics nationallicence Moore Vicky Centre for Extremophile Research, Department of Biology and Biochemistry, University of Bath, BA2 7AY, Bath, UK aut Kanu Ami Centre for Extremophile Research, Department of Biology and Biochemistry, University of Bath, BA2 7AY, Bath, UK aut Byron Olwyn FBLS Division of Infection and Immunity, Glasgow Biomedical Research Centre, University of Glasgow, G12 8TA, Glasgow, UK aut Campbell Gordon FBLS Division of Infection and Immunity, Glasgow Biomedical Research Centre, University of Glasgow, G12 8TA, Glasgow, UK aut Danson Michael Centre for Extremophile Research, Department of Biology and Biochemistry, University of Bath, BA2 7AY, Bath, UK aut Hough David Centre for Extremophile Research, Department of Biology and Biochemistry, University of Bath, BA2 7AY, Bath, UK aut Crennell Susan Centre for Extremophile Research, Department of Biology and Biochemistry, University of Bath, BA2 7AY, Bath, UK aut Extremophiles Springer Japan 15/3(2011-05-01), 327-336 1431-0651 15:3<327 2011 15 792 https://doi.org/10.1007/s00792-011-0363-6 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s00792-011-0363-6 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Moore Vicky Centre for Extremophile Research, Department of Biology and Biochemistry, University of Bath, BA2 7AY, Bath, UK aut NATIONALLICENCE 700 1- Kanu Ami Centre for Extremophile Research, Department of Biology and Biochemistry, University of Bath, BA2 7AY, Bath, UK aut NATIONALLICENCE 700 1- Byron Olwyn FBLS Division of Infection and Immunity, Glasgow Biomedical Research Centre, University of Glasgow, G12 8TA, Glasgow, UK aut NATIONALLICENCE 700 1- Campbell Gordon FBLS Division of Infection and Immunity, Glasgow Biomedical Research Centre, University of Glasgow, G12 8TA, Glasgow, UK aut NATIONALLICENCE 700 1- Danson Michael Centre for Extremophile Research, Department of Biology and Biochemistry, University of Bath, BA2 7AY, Bath, UK aut NATIONALLICENCE 700 1- Hough David Centre for Extremophile Research, Department of Biology and Biochemistry, University of Bath, BA2 7AY, Bath, UK aut NATIONALLICENCE 700 1- Crennell Susan Centre for Extremophile Research, Department of Biology and Biochemistry, University of Bath, BA2 7AY, Bath, UK aut NATIONALLICENCE 773 0- Extremophiles Springer Japan 15/3(2011-05-01), 327-336 1431-0651 15:3<327 2011 15 792 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer