Secondary Structure of Recombinant Human Cystathionine β-Synthase in Aqueous Solution: Effect of Ligand Binding and Proteolytic Truncation

The secondary structural composition and substrate-induced conformational changes of recombinant human cystathionine β-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 β-sheet (1689, 1638, and 1627 cm−1), α-helix (1658 cm−1), β-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% β-sheet, 25–30% α-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 Garnierʹs method shows that human CBS may contain a β-sheet/α-helix/β-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.