Protective effect of disaccharides on restriction endonucleases during drying under vacuum.
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 lambda DNA-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 degrees C, but it shows reduced activity after 30-day storage at either 4 degrees 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 degrees C and for 7 days at room temperature.
Uritani
M
M
Department of Chemistry, Faculty of Science, Shizuoka University.
Takai
M
M
Yoshinaga
K
K
D004262:DNA Restriction Enzymes / Q000737:chemistry* / Q000378:metabolism; D015245:Deoxyribonuclease BamHI / Q000737:chemistry / Q000378:metabolism; D015246:Deoxyribonuclease EcoRI / Q000737:chemistry / Q000378:metabolism; D015247:Deoxyribonuclease HindIII / Q000737:chemistry; D003890:Desiccation; D004187:Disaccharides / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D004795:Enzyme Stability; D008320:Maltose / Q000494:pharmacology; D013395:Sucrose / Q000494:pharmacology; D013696:Temperature; D013997:Time Factors; D014199:Trehalose / Q000494:pharmacology; D014618:Vacuum
D004187:Disaccharides; D013395:Sucrose; D008320:Maltose; D014199:Trehalose; D004262:DNA Restriction Enzymes; D015245:Deoxyribonuclease BamHI; D015246:Deoxyribonuclease EcoRI; D015247:Deoxyribonuclease HindIII