Organic solvent binding to crystalline subtilisin1 in mostly aqueous media and in the neat solvents.
The X-ray crystal structures of the protease subtilisin Carlsberg in 40% acetonitrile and in 20% dioxane have been determined to at least 2.3 A resolution, and their solvent binding patterns have been compared to those observed in the neat organic solvents. The structures of the protein in the two aqueous-organic mixtures are essentially the same as in pure water, acetonitrile, and dioxane. Interestingly, the enzyme-bound organic solvent molecules tend to congregate in the active site. Three of the five bound acetonitrile molecules observed in the structure of subtilisin in 40% acetonitrile are situated in the enzyme active site, as is the single enzyme-bound dioxane molecule observed in 20% dioxane (whose location is distinct from that of any bound acetonitrile molecule). Furthermore, the organic solvent molecules detected in the enzyme active site in the aqueous-organic mixtures are in the same locations as in the structures in the corresponding neat organic solvents.
Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts, 02139, USA.
J L
JL
Schmitke
L J
LJ
Stern
A M
AM
Klibanov
D000097:Acetonitriles / Q000378:metabolism; D001407:Bacillus / Q000201:enzymology; D001426:Bacterial Proteins / Q000737:chemistry; D001665:Binding Sites; D018360:Crystallography, X-Ray; D004146:Dioxanes / Q000378:metabolism; D010450:Endopeptidases / Q000737:chemistry; D008958:Models, Molecular; D011485:Protein Binding / Q000502:physiology; D012997:Solvents / Q000378:metabolism*; D013381:Subtilisins / Q000737:chemistry*
D000097:Acetonitriles; D001426:Bacterial Proteins; D004146:Dioxanes; D012997:Solvents; D010450:Endopeptidases; D013381:Subtilisins; C025223:1,4-dioxane; C032159:acetonitrile