title: Analysis of the conformational stability of the active domain of recombinant mouse TIMP-1 by intrinsic fluorescence.
abstract: Intrinsic fluorescence was used to examine the stability of an active, N-terminal domain of mouse tissue inhibitor of metalloproteinase (TIMP-1) fused with an N-terminal polyhistidine tag. Emission and quenching studies suggested that the single tryptophan is on the protein surface partially exposed to solvent. The TIMP-1 recombinant unfolded reversibly in the presence of guanidinium chloride with the transition midpoint at 2.35M; extrapolation gave a stabilization free energy of 5.1 kcal mol-1 at 25 degrees C. Analysis of the temperature dependence of the fluorescence intensity gave a melting transition with midpoint at 51 degrees C and an enthalpy and heat capacity change on unfolding of 32 kcal mol-1 and 0.45 kcal K-1 mol-1, respectively, values comparable to other single domain proteins. Comparison with literature data indicated that the stability of mouse recombinant TIMP-1 more closely resembled that of human metalloproteinase inhibitor TIMP-2 than TIMP-1 despite closer homology to the human TIMP-1 protein.
authors.affiliation: Division of Life Sciences, Rutgers University, Piscataway, New Jersey 08855-1059, USA.
authors.forename: S S
authors.initials: SS
authors.lastname: Rajan
authors.forename: D T
authors.initials: DT
authors.lastname: Denhardt
authors.forename: H
authors.initials: H
authors.lastname: Lackland
authors.forename: R D
authors.initials: RD
authors.lastname: Ludescher
mesh_terms: D020106:Acrylamide; D000178:Acrylamides / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D019791:Guanidine / Q000494:pharmacology; D051379:Mice; D011487:Protein Conformation; D011489:Protein Denaturation / Q000187:drug effects; D017510:Protein Folding; D011994:Recombinant Proteins / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013696:Temperature; D013816:Thermodynamics; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000737:chemistry / Q000378:metabolism*; D014364:Tryptophan / Q000737:chemistry / Q000378:metabolism
chemical_list: D000178:Acrylamides; D011994:Recombinant Proteins; D019715:Tissue Inhibitor of Metalloproteinase-1; D020106:Acrylamide; D014364:Tryptophan; D019791:Guanidine