title: Purification, characterization, and crystallization of alliinase from garlic.
abstract: Glycosylated dimeric alliinase (EC 4.4.1.4) was purified to homogeneity from its natural source, garlic. With 660 units/mg, the specific enzymatic activity of the pure enzyme is the highest reported to date. Based on both CD spectroscopy data and sequence-derived secondary structure prediction, the alpha-helix content of alliinase was estimated to be about 30%. Comparisons of all available amino acid sequences of alliinases revealed a common cysteine pattern of the type C-x18-19-C-x-C-x2-C-x5-C-x6-C in the N-terminal part of the sequences. This pattern is conserved in alliinases but absent in other pyridoxal 5'-phosphate-dependent enzymes. It suggests the presence of an epidermal growth factor-like domain in the three-dimensional structures of alliinases, making them unique among the various families of pyridoxal 5'-phosphate-dependent enzymes. Well-ordered three-dimensional crystals of garlic alliinase were obtained in four different forms. The best diffraction was observed with crystal form IV (space group P2(1)2(1)2(1), a=68.4, b=101.1, c=155.7 A) grown from an ammonium sulfate solution. These crystals diffract to at least 1.5 A resolution at a synchrotron source and are suitable for structure determination.
authors.affiliation: Department of Structural Biology and Crystallography, Institute of Molecular Biotechnology, Beutenbergstrasse 11, D-07745 Jena, Germany.
authors.forename: E Bartholomeus
authors.initials: EB
authors.lastname: Kuettner
authors.forename: Rolf
authors.initials: R
authors.lastname: Hilgenfeld
authors.forename: Manfred S
authors.initials: MS
authors.lastname: Weiss
mesh_terms: D000595:Amino Acid Sequence; D013437:Carbon-Sulfur Lyases / Q000737:chemistry* / Q000302:isolation & purification; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D004815:Epidermal Growth Factor / Q000737:chemistry; D005737:Garlic / Q000737:chemistry*; D007700:Kinetics; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D013056:Spectrophotometry, Ultraviolet
chemical_list: D004815:Epidermal Growth Factor; D013437:Carbon-Sulfur Lyases; C019243:alliin lyase