beta-glycosidase from the hyperthermophilic archaeon Sulfolobus solfataricus: structure and activity in the presence of alcohols.
beta-Glycosidase from the extreme thermophilic archaeon Sulfolobus solfataricus is a tetrameric protein with a molecular mass of 240 kDa, stable in the presence of detergents, and with a maximal activity at temperatures above 95 degrees C. Understanding the structure-activity relationships of the enzyme under different conditions is of fundamental importance for both theoretical and applicative purposes. In this paper we report the effect of methanol, ethanol, 1-propanol, and 1-butanol on the activity of S. solfataricus beta-glycosidase expressed in Escherichia coli. The alcohols stimulated the enzyme activity, with 1-butanol producing its maximum effect at a lower concentration than the other alcohols. The structure of the enzyme was studied in the presence of 1-butanol by circular dichroism, and Fourier-transform infrared and fluorescence spectroscopies. Circular dichroism and steady-state fluorescence measurements revealed that at low temperatures the presence of the alcohol produced no significant changes in the tertiary structure of the enzyme. However, time-resolved fluorescence data showed that the alcohol modifies the protein microenvironment, leading to a more flexible enzyme structure, which is probably responsible for the enhanced enzymatic activity.
D'Auria
S
S
Istituto di Biochimica delle Proteine ed Enzimologia, C.N.R., Via Marconi, 10 80125 Naples, Italy. dauria@dafne.ibpe.na.cnr.it
Nucci
R
R
Rossi
M
M
Bertoli
E
E
Tanfani
F
F
Gryczynski
I
I
Malak
H
H
Lakowicz
J R
JR
D020001:1-Butanol / Q000737:chemistry*; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D011489:Protein Denaturation; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet; D013329:Structure-Activity Relationship; D016944:Sulfolobus / Q000201:enzymology*
D020001:1-Butanol; D006026:Glycoside Hydrolases