Conformational stability of 17 beta-hydroxysteroid dehydrogenase from the fungus Cochliobolus lunatus.
The functional activities of proteins are closely related to their molecular structure and understanding their structure-function relationships remains one of the intriguing problems of molecular biology. We investigated structural changes in 17beta-hydroxysteroid dehydrogenase from the fungus Cochliobolus lunatus (17beta-HSDcl) induced by pH, temperature, salt, urea, guanidine hydrochloride, and coenzyme NADPH binding. At 25 degrees C and within the relatively narrow pH range of 7.0-9.0, 17beta-HSDcl exists in its native conformation as a dimer. This native conformation is thermally stable up to 40 degrees C in this pH range. At 25 degrees C and pH 2.0 in the presence of 150-300 mM NaCl, 17beta-HSDcl forms soluble aggregates enriched in alpha-helical and beta-sheet structures. At higher temperatures and NaCl concentrations, these soluble aggregates start to precipitate. The denaturants urea and guanidine hydrochloride unfold 17beta-HSDcl at concentrations of 1.2 and 0.4 M, respectively. Binding of the coenzyme NADPH to 17beta-HSDcl causes local structural changes that do not significantly affect the thermal stability of this protein.
Ulrih
Natasa Poklar
NP
Department of Food Science and Technology, University of Ljubljana, Slovenia. natasa.poklar@bf.uni-lj.si
Lanisnik Rizner
Tea
T
D015067:17-Hydroxysteroid Dehydrogenases / Q000737:chemistry* / Q000378:metabolism*; D001203:Ascomycota / Q000201:enzymology*; D002942:Circular Dichroism; D004795:Enzyme Stability; D019791:Guanidine; D006863:Hydrogen-Ion Concentration; D011487:Protein Conformation; D011489:Protein Denaturation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013056:Spectrophotometry, Ultraviolet; D013696:Temperature; D014508:Urea
D011994:Recombinant Proteins; D014508:Urea; D015067:17-Hydroxysteroid Dehydrogenases; C044256:3 (or 17)-beta-hydroxysteroid dehydrogenase; D019791:Guanidine