Effect of glycosylation on the mechanism of renaturation of invertase from yeast

• Published in: The Journal of biological chemistry Vol. 263; no. 18; pp. 8832 - 8837
• Main Authors: Schülke, N, Schmid, F X
• Format: Journal Article
• Language: English
• Published: Bethesda, MD Elsevier Inc 25.06.1988; American Society for Biochemistry and Molecular Biology
• Subjects: ACTIVIDAD ENZIMATICA; ACTIVITE ENZYMATIQUE; beta-Fructofuranosidase; Biological and medical sciences; Circular Dichroism; ENZYMIC ACTIVITY; FRUCTOFURANOSIDASA; FRUCTOFURANOSIDASE; Fundamental and applied biological sciences. Psychology; Glycoside Hydrolases - metabolism; Glycosylation; Growth, nutrition, metabolism, transports, enzymes. Molecular biology; Hydrogen-Ion Concentration; Isoenzymes - metabolism; Kinetics; LEVADURA; LEVURE; Microbiology; Molecular Weight; Mycology; Protein Conformation; Protein Denaturation; PROTEINAS; PROTEINE; PROTEINS; Saccharomyces cerevisiae; Saccharomyces cerevisiae - enzymology; Spectrometry, Fluorescence; YEASTS; Fungi; Denaturation renaturation; Yeast; Stability; β-D-Fructofuranosidase; Ascomycetes; Glycosylation; Property structure relationship; Saccharomyces cerevisiae; Thallophyta
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)68382-6

N-Glycosylation occurs cotranslationally as soon as the growing polypeptide chain enters the endoplasmic reticulum, before the final native-like folded state is reached. We examined the role of the carbohydrate chains in the mechanism of protein folding. The in vitro folding and association of yeast invertase are used as an experimental system. External invertase contains approximately 50% carbohydrate, whereas cytoplasmic invertase is not glycosylated. The functional native state of both proteins is a homodimer. At pH greater than or equal to 6.5 and at protein concentrations below 3 micrograms/ml, the kinetics of reactivation and the final yields are similar for the two invertases. For both proteins, reactivation is a sequential reaction with a lag phase at the beginning. The nonglycosylated protein tends to aggregate during reactivation at low pH and at protein concentrations above 3 micrograms/ml. After separation of inactive material, the renatured protein is indistinguishable from the original native state by a number of physicochemical and functional criteria. The results suggest that the carbohydrate moiety does not affect the mechanism of folding and association of invertase. However, glycosylation improves the solubility of unfolded or partially folded invertase molecules and hence leads to a suppression of irreversible aggregation. Such a protective effect may also be important for the in vivo maturation of nascent glycosylated protein chains.