Protective Effect of Disaccharides on Restriction Endonucleases during Drying under Vacuum

• Published in: Journal of biochemistry (Tokyo) 1995-04, Vol.117 (4), p.774-779
• Main Authors: Uritani, Masahiro, Takai, Megumi, Yoshinaga, Koichi
• Format: Article
• Language: English
• Subjects: Deoxyribonuclease BamHI - chemistry; Deoxyribonuclease BamHI - metabolism; Deoxyribonuclease EcoRI - chemistry; Deoxyribonuclease EcoRI - metabolism; Deoxyribonuclease HindIII - chemistry; Desiccation; disaccharide; Disaccharides - pharmacology; DNA Restriction Enzymes - chemistry; DNA Restriction Enzymes - metabolism; drying; endonuclease; enzyme; Enzyme Activation - drug effects; Enzyme Stability; Maltose - pharmacology; Sucrose - pharmacology; Temperature; Time Factors; trehalose; Trehalose - pharmacology; Vacuum
• ISSN: 0021-924X
• DOI: 10.1093/oxfordjournals.jbchem.a124775

Desiccation by vacuum-drying inactivates the restriction endonuclease HindIII completely. However, when dried in the presence of a disaccharide such as trehalose, maltose, or sucrose, the endonuclease retains its λDNA-cleaving activity and produces the same digestive fragments as does the intact enzyme. Thus, the disaccharides are effective in protecting the restriction enzyme in terms of both recognition and accurate cleavage of the substrate. Among the disaccharides, trehalose protects the enzyme most effectively; and it also stabilizes the enzyme during dilution in aqueous solution. The restriction enzyme dried with trehalose maintains its activity without detectable loss for at least 4 days at 37°C, but it shows reduced activity after 30-day storage at either 4°C or room temperature. Trehalose also protects other restriction endonucleases, EcoRI and BamHI, from inactivation during vacuum-drying, whereas drying them alone leads to severe loss of their activity. The restriction endonucleases dried with trehalose retain their activities for at least 20 days at 4°C and for 7 days at room temperature