A comparative study of HPr proteins from extremophilic organisms

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Title: A comparative study of HPr proteins from extremophilic organisms
Author: Syed Ali, Abbas Razvi
Abstract: A thermodynamic study of five homologous HPr proteins derived from organisms inhabiting diverse environments has been undertaken . The aim of this study was to further our understanding of protein stabilization in extremes of environment . Two of the proteins were derived from moderate thermophiles (Streptococcus thermophilus and Bacillus staerothermophilus ) and two from haloalkaliphilic organisms (Bacillus halodurans and Oceanobacillus iheyensis ) ; these proteins were compared with HPr from the mesophile Bacillus subtilus . Genes for three of these homologous HPr proteins were for the first time cloned from their respective organisms into expression vectors and they were over -expressed and purified in Escherichia coli . Stability measurements were performed on these proteins under a variety of solution conditions (varying pH , salinity and temperature ) by thermal and solvent induced denaturation experiments . Stability curves were determined for every homologue and these reveal very similar conformational stability for these homologues at their habitat temperatures . The BstHPr homologue is the most thermostable and also has the highest G25 ; the stability of other homologues was ranked as Bst >Bh >St >Bs >OiHPr . Other key thermodynamic parameters , like Cp , have been estimated for all the homologues and it was found that these values are identical within errors of estimation . Also , it was found that the values of TS are very similar for these homologues . Together these observations allow us to propose a thermodynamic mechanism toward achieving higher Tm . The crystal structures of the BstHPr and a single tryptophan -containing variant (BstF29W ) of this homologue are also reported here . Also reported is a domain -swapped dimeric structure for the BstF29W variant , together with a detailed investigation into the solution oligomeric nature of this protein . The crystal structure of BstHPr is analyzed to enumerate various stabilizating interactions like hydrogen bonds and salt -bridges and these were compared with those for the mesophilic homologue BsHPr . Finally , an analysis of sequence alignments together with structural information for these homologues has allowed design of numerous variants of both Bs and BstHPr . A detailed thermodynamic study of these variants is presented in an attempt to understand the origins of the differences in stability of the HPr homologues .
URI: http : / /hdl .handle .net /1969 .1 /3163
Date: 2006-04-12

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A comparative study of HPr proteins from extremophilic organisms. Available electronically from http : / /hdl .handle .net /1969 .1 /3163 .

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