title: Effect of chemical modifications on freeze denaturation of lactate dehydrogenase.
abstract: Lactate dehydrogenase (LDH) was chemically ethyl-acetimidated (EA-), dimethyladipimidated (DMA-), carbamylated, acetylated, acetoacetylated, or succinylated in order to alter the ionic charges on the epsilon-amino group of lysine residues. Acetylation, acetoacetylation, and succinylation, which change the positive charge at the lysine side chains to a negative one, inactivated the enzymic activity, but the rest of the modifications exerted no such inactivating effects. The active modified enzymes were subjected to freeze denaturation study, using the enzymic activity as an indication of the degree of the denaturation. The active enzymes were diluted with deionized water and stored in a freezer (-23 degrees C) for 1-3 days. Enzymic activity was assayed immediately after thawing. All the modified enzymes retained their activity even after the 3-day frozen storage, while the control or native enzyme lost its activity within 1 day of storage. Furthermore, the modified LDHs freeze-stored in 0.2 M monosodium glutamate (MSG) or 0.2 M lysine-hydrochloride (Lys-HCl) retained their activity. The cryoprotective effects exerted by the modifications and by 0.2 M MSG seemed to be synergistic, whereas those exerted by the modifications and by 0.2 M Lys-HCl did not. The mechanisms of cryoprotection and freeze denaturation are discussed in relationship with the cryoprotective effect exerted by already known cryoprotectants, such as sucrose or dimethyl sulfoxide.
authors.affiliation: Department of Chemistry, Faculty of Science and Technology, Sophia University, Tokyo, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Seguro
authors.forename: T
authors.initials: T
authors.lastname: Tamiya
authors.forename: T
authors.initials: T
authors.lastname: Tsuchiya
authors.forename: J J
authors.initials: JJ
authors.lastname: Matsumoto
mesh_terms: D000818:Animals; D003451:Cryoprotective Agents; D004789:Enzyme Activation; D005615:Freezing; D007202:Indicators and Reagents; D007770:L-Lactate Dehydrogenase; D013329:Structure-Activity Relationship; D013552:Swine
chemical_list: D003451:Cryoprotective Agents; D007202:Indicators and Reagents; D007770:L-Lactate Dehydrogenase