title: A novel membrane-anchored cytochrome c-550 of alkaliphilic Bacillus clarkii K24-1U: expression, molecular features and properties of redox potential.
abstract: A membrane-anchored cytochrome c-550, which is highly expressed in obligately alkaliphilic Bacillus clarkii K24-1U, was purified and characterized. The protein contained a conspicuous sequence of Gly(22)-Asn(34), in comparison with the other Bacillus small cytochromes c. Analytical data indicated that the original and lipase-treated intermediate forms of cytochrome c-550 bind to fatty acids of C(15), C(16) and C(17) chain lengths and C(15) chain length, respectively, and it was considered that these fatty acids are bound to glycerol-Cys(18). Since there was a possibility that the presence of a diacylglycerol anchor contributed to the formation of dimeric states of this protein (20 and 17 kDa in SDS-PAGE), a C18M (Cys(18) --> Met)-cytochrome c-550 was constructed. The molecular mass of the C18M-cytochrome c-550 was determined as 15 and 10 kDa in SDS-PAGE and 23 kDa in blue native PAGE. The C18M-cytochrome c-550 bound with or without Triton X-100 formed a tetramer as the original cytochrome c-550 bound with Triton X-100, as determined by gel filtration. The midpoint redox potential of cytochrome c-550 as determined by redox titration was +83 mV, while that determined by cyclic voltammetric measurement was +7 mV. The above results indicate that cytochrome c-550 is a novel cytochrome c.
authors.affiliation: Research Institute of Genome-based Biofactory, National Institute of Advanced Industrial Science and Technology (AIST), Tsukisamu-Higashi, Toyohira-ku, Sapporo, 062-8517, Japan.
authors.forename: Shinichi
authors.initials: S
authors.lastname: Ogami
authors.forename: Shoichi
authors.initials: S
authors.lastname: Hijikata
authors.forename: Tamotsu
authors.initials: T
authors.lastname: Tsukahara
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Mie
authors.forename: Toshihide
authors.initials: T
authors.lastname: Matsuno
authors.forename: Naoki
authors.initials: N
authors.lastname: Morita
authors.forename: Isao
authors.initials: I
authors.lastname: Hara
authors.forename: Koji
authors.initials: K
authors.lastname: Yamazaki
authors.forename: Norio
authors.initials: N
authors.lastname: Inoue
authors.forename: Atsushi
authors.initials: A
authors.lastname: Yokota
authors.forename: Tamotsu
authors.initials: T
authors.lastname: Hoshino
authors.forename: Kazuaki
authors.initials: K
authors.lastname: Yoshimune
authors.forename: Isao
authors.initials: I
authors.lastname: Yumoto
mesh_terms: D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology*; D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D003574:Cytochrome c Group / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D005227:Fatty Acids / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013056:Spectrophotometry, Ultraviolet
chemical_list: D003574:Cytochrome c Group; D017931:DNA Primers; D005227:Fatty Acids; D008565:Membrane Proteins; C031732:cytochrome C-550