title: Functional analyses of carnivorous plant-specific amino acid residues in S-like ribonucleases.
abstract: Unlike plants with no carnivory, carnivorous plants seem to use S-like ribonucleases (RNases) as an enzyme for carnivory. Carnivorous plant-specific conserved amino acid residues are present at four positions around the conserved active site (CAS). The roles of these conserved amino acid residues in the enzymatic function were explored in the current study by preparing five recombinant variants of DA-I, the S-like RNase of Drosera adelae. The kcat and kcat/Km values of the enzymes revealed that among the four variants with a single mutation, the serine to glycine mutation at position 111 most negatively influenced the enzymatic activity. The change in the bulkiness of the amino acid residue side-chain seemed to be the major cause of the above effect. Modeling of the three dimensional (3D) structures strongly suggested that the S to G mutation at 111 greatly altered the overall enzyme conformation. The conserved four amino acid residues are likely to function in keeping the two histidine residues, which are essential for the cleavage of RNA strands, and the CAS in the most functional enzymatic conformation.
authors.affiliation: Major in Integrative Bioscience and Biomedical Engineering, Graduate School of Science and Engineering, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480, Japan.
authors.forename: Naoki
authors.initials: N
authors.lastname: Arai
authors.affiliation: Major in Integrative Bioscience and Biomedical Engineering, Graduate School of Science and Engineering, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480, Japan.
authors.forename: Emi
authors.initials: E
authors.lastname: Nishimura
authors.affiliation: Major in Integrative Bioscience and Biomedical Engineering, Graduate School of Science and Engineering, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480, Japan.
authors.forename: Yo
authors.initials: Y
authors.lastname: Kikuchi
authors.affiliation: Major in Integrative Bioscience and Biomedical Engineering, Graduate School of Science and Engineering, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480, Japan; Department of Biology, Faculty of Education and Integrated Arts and Sciences, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480, Japan. Electronic address: ohyama@waseda.jp.
authors.forename: Takashi
authors.initials: T
authors.lastname: Ohyama
mesh_terms: D000595:Amino Acid Sequence; D060435:Carnivory / Q000502:physiology; D029787:Drosera / Q000737:chemistry* / Q000201:enzymology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012260:Ribonucleases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013329:Structure-Activity Relationship
chemical_list: D010940:Plant Proteins; D011994:Recombinant Proteins; D012260:Ribonucleases; C489908:S-like RNase protein, plant
keywords: Carnivorous plants; Drosera adelae; Protein structure; S-like ribonuclease; k(cat)/K(m)