Unique oligomeric intermediates of bovine liver catalase.
Catalases, although synthesized from single genes and built up from only one type of subunit, exist in heterogeneous form with respect to their conformations and association states in biological systems. This heterogeneity is not of genetic origin, but rather reflects the instability of this oligomeric heme enzyme. To understand better the factors that stabilize the various association states of catalase, we performed studies on the multimeric intermediates that are stabilized during guanidine-hydrochloride- and urea-induced unfolding of bovine liver catalase (BLC). For the first time, we have observed an enzymatically active, folded dimer of native BLC. This dimer has slightly higher enzymatic activity and altered structural properties compared to the native tetramer. Comparative studies of the effect of NaCl, GdmCl, and urea on BLC show that cation binding to negatively charged groups present in amino acid side chains of the enzyme leads to stabilization of an enzymatically active, folded dimer of BLC. Besides the folded dimer, an enzymatically active expanded tetramer and a partially unfolded, enzymatically inactive dimer of BLC were also observed. A complete recovery of native enzyme was observed on refolding of expanded tetramers and folded dimers; however, a very low recovery (maximum of approximately 5%) of native enzyme was observed on refolding of partially unfolded dimers and fully unfolded monomers.
Prakash
Koodathingal
K
Molecular and Structural Biology Division, Central Drug Research Institute, Lucknow 226 001, India.
Prajapati
Shashi
S
Ahmad
Atta
A
Jain
S K
SK
Bhakuni
Vinod
V
D000817:Anilino Naphthalenesulfonates / Q000494:pharmacology; D000818:Animals; D002374:Catalase / Q000737:chemistry*; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D003432:Cross-Linking Reagents / Q000494:pharmacology; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D005456:Fluorescent Dyes / Q000494:pharmacology; D005976:Glutaral / Q000737:chemistry / Q000494:pharmacology; D019791:Guanidine / Q000494:pharmacology; D008099:Liver / Q000201:enzymology*; D017510:Protein Folding; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D012965:Sodium Chloride / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D013997:Time Factors; D014508:Urea / Q000494:pharmacology
D000817:Anilino Naphthalenesulfonates; D003432:Cross-Linking Reagents; D005456:Fluorescent Dyes; D012965:Sodium Chloride; C027132:1-anilino-8-naphthalenesulfonate; D014508:Urea; D002374:Catalase; D019791:Guanidine; D005976:Glutaral