title: Secondary structure of recombinant human cystathionine beta-synthase in aqueous solution: effect of ligand binding and proteolytic truncation.
abstract: The secondary structural composition and substrate-induced conformational changes of recombinant human cystathionine beta-synthase (CBS) in aqueous solution have been investigated in its full-length form (tetramer of 63-kDa subunits) by Fourier transform infrared (FT-IR) and circular dichroism (CD) spectroscopies. In addition, structural comparison of a proteolytic truncated form (dimer of 45-kDa subunits) to that of the full-length enzyme has also been carried out. Second-derivative and Fourier self-deconvolutional enhanced infrared spectra revealed amide I band components ascribed to beta-sheet (1689, 1638, and 1627 cm(-1)), alpha-helix (1658 cm(-1)), beta-turn (1679 and 1668 cm(-1)), and unordered (1651 cm(-1)) structures in the spectra of the full-length enzyme. Quantitative analysis of FT-IR and CD spectra reveals that the full-length enzyme consists of about 48-53% beta-sheet, 25-30% alpha-helix, 8-10% turn, and 10-19% unordered structures. Under constraint of the spectroscopic data, theoretical prediction of locations of these secondary structural elements using Garnier's method shows that human CBS may contain a beta-sheet/alpha-helix/beta-sheet core structure. Second-derivative spectrum of the truncated enzyme exhibited all the major spectral features that are present in the full-length enzyme, indicating a preservation of the core structure of the enzyme. Significant differences were observed between the infrared spectra of the enzymes with or without the substrate, serine, indicating a substrate-induced conformational change in the enzyme, which did not result in a change in overall composition of secondary structural content based on quantitative analysis of FT-IR and far-UV CD spectra.
authors.affiliation: Department of Pharmaceutical Sciences, School of Pharmacy, University of Colorado Health Sciences Center, Denver 80262, USA.
authors.forename: A
authors.initials: A
authors.lastname: Dong
authors.forename: V
authors.initials: V
authors.lastname: Kery
authors.forename: J
authors.initials: J
authors.lastname: Matsuura
authors.forename: M C
authors.initials: MC
authors.lastname: Manning
authors.forename: J P
authors.initials: JP
authors.lastname: Kraus
authors.forename: J F
authors.initials: JF
authors.lastname: Carpenter
mesh_terms: D000595:Amino Acid Sequence; D002942:Circular Dichroism; D003541:Cystathionine beta-Synthase / Q000737:chemistry* / Q000378:metabolism; D019281:Dimerization; D006801:Humans; D008024:Ligands; D008969:Molecular Sequence Data; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017550:Spectroscopy, Fourier Transform Infrared; D014357:Trypsin / Q000378:metabolism
chemical_list: D008024:Ligands; D011994:Recombinant Proteins; D014357:Trypsin; D003541:Cystathionine beta-Synthase