Invertase from a strain of Rhodotorula glutinis.
An invertase (beta-D-fructofuranoside fructohydrolase, EC 3.2.1.26) from Rhodotorula glutinis was purified by ammonium sulfate fractionation, gel filtration and anion exchange chromatography. Invertase molecular weight was estimated to be 100 kDa by analytical gel filtration and 47 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Molecular mass determinations indicated that the native enzyme exists as a homodimer. It is a glycoprotein that contains 19% carbohydrate. The enzyme attacks beta-D-fructofuranoside (raffinose, stachyose and sucrose) from the fructose end. It has a K(m) of 0.227 M and a V(max) of 0.096 micromol/min with sucrose as a substrate. Invertase activity is stable between pH 2.6 and 5.5 for 30 min, maximum activity being observed at pH 4.5. The activation energy was 6520 cal/mol. The enzyme is stable between 20 and 60 degrees C. Mg(2+) and Ca(2+) ions stimulated invertase activity 3-fold, while Fe(2+), K(+), Co(2+), Na(+) and Cu(2+) increased activity about 2-fold. The transfructosylation reaction could not be observed. This enzyme is of particular interest since it appears to have a high hydrolytic activity in 1 M sucrose solution. This fact would make the enzymatic hydrolysis process economically efficient for syrup production using by-products with high salt and sugar contents such as sugar cane molasses.
Rubio
María C
MC
Department of Industrial Microbiology, Institute of Biotechnology, Faculty of Biochemistry, Chemistry and Pharmacy, University of Tucumán, Ayacucho 491-4000, San Miguel de Tucumán, Argentina. biotec@unt.edu.ar
Runco
Rosa
R
Navarro
Antonio R
AR
D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008670:Metals / Q000378:metabolism; D008970:Molecular Weight; D012248:Rhodotorula / Q000201:enzymology*; D013379:Substrate Specificity; D013696:Temperature; D043324:beta-Fructofuranosidase
D008670:Metals; D006026:Glycoside Hydrolases; D043324:beta-Fructofuranosidase