Purification and molecular properties of rabbit lung indolamine N-methyltransferase.
Indolamine N-methyltransferase (INMT) has been purified to an apparent homogeneity from rabbit lung, and some of its catalytic and physicochemical properties have been examined. The enzyme is a monomeric protein with a molecular weight of 31,500 +/- 1000, a molecular Stokes radius of 21.5 A, and a diffusion coefficient of 8.7 X 10(-7) cm2/s. The frictional ratio of the native enzyme (1.05) suggests that the shape of the molecule is nearly spherical. Denaturation experiments performed with increasing concentrations of guanidine hydrochloride (Gdn-HCl) at neural pH indicated that the active site of the enzyme was destroyed by a structural rearrangement of the protein molecule without large change in its size and shape. The final state reached in 6.0 M Gdn . HCl seemed to correspond to a disulfide cross-linked randomly coiled polypeptide. Full normalization of the fluorescent parameter was attained only in the presence of 0.1 M beta-mercaptoethanol. A structural rearrangement has been observed upon acidification of INMT from pH 7.0 to pH 2.0. At pH 4.5, most of the peptide backbone appeared to be unorganized, but further acidification to pH 2.0 produced a reorganization of protein structure which became able to bind 8-anilino-1-naphthalenesulfonate. The data support the hypothesis that the enzyme structure results from the close package of organized regions joined by structureless segments.
D000818:Animals; D055598:Chemical Phenomena; D002621:Chemistry; D002850:Chromatography, Gel; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D005454:Fluorescence Polarization; D007700:Kinetics; D008168:Lung / Q000201:enzymology*; D008780:Methyltransferases / Q000302:isolation & purification*; D008970:Molecular Weight; D011817:Rabbits; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet; D014363:Tryptamines / Q000302:isolation & purification
D014363:Tryptamines; D008780:Methyltransferases; C021438:indoleamine-N-methyltransferase