Determination of the secondary structure of Kluyveromyces lactis beta-galactosidase by circular dichroism and its structure-activity relationship as a function of the pH.
The secondary structure of Kluyveromyces lactis beta-galactosidase was determined by circular dichroism. It is mainly a beta-type protein, having 22% beta-turns, 14% parallel beta-sheet, 25% antiparallel beta-sheet, 34% unordered structure, and only 5% alpha-helix. The structure-activity relationship as a function of the pH was also studied. The pH conditions leading to the highest secondary structure content (100% ellipticity) of the enzyme was found at pH 7.0; at pH 6.5-7.0, the percent ellipticity decreased slightly, suggesting little structural change, but the activity decreased significantly, probably because of variations in critical residues. On the other hand, at pH's above 7.0, a more noticeable change in ellipticity was observed due to structural changes; the CD analysis showed a small increase in the helical content toward higher pH, whereas the maximum activity was found at pH 7.5, meaning that the changes produced in the secondary structure at this pH favored the interaction between the enzyme and the substrate.
Tello-Solís
Salvador R
SR
Area de Biofisicoquímica, Departamento de Química, Universidad Autónoma Metropolitana, Iztapalapa. Avenida San Rafael Atlixco 186, Col. Vicentina, Mexico DF 09340, Mexico. srts@xanum.uam.mx
Jiménez-Guzmán
Judith
J
Sarabia-Leos
Christian
C
Gómez-Ruíz
Lorena
L
Cruz-Guerrero
Alma E
AE
Rodríguez-Serrano
Gabriela M
GM
García-Garibay
Mariano
M
D002850:Chromatography, Gel; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D005656:Fungal Proteins / Q000096:biosynthesis / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007716:Kluyveromyces / Q000201:enzymology*; D043322:Lactase / Q000737:chemistry / Q000302:isolation & purification; D011487:Protein Conformation; D013329:Structure-Activity Relationship; D001616:beta-Galactosidase / Q000737:chemistry*
D005656:Fungal Proteins; D043322:Lactase; D001616:beta-Galactosidase