Misa Masanari

Hiroshima University, Hirosima, Hiroshima, Japan

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Publications (6)12.65 Total impact

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    ABSTRACT: Apo-cytochomes c without heme are usually unstructured. Here we showed that apo-form of thermophilic Hydrogenophilus thermoluteolus cytochrome c' (PHCP) was a monomeric protein with high helix content. Apo-PHCP was thermally stable, possibly due to the hydrophobic residues and ion pairs. PHCP is the first example of a structured apo-cytochrome c', which will expand our view of hemoprotein structure formation.
    Bioscience Biotechnology and Biochemistry 07/2014; 78(7):1191-1194. · 1.27 Impact Factor
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    ABSTRACT: Shewanella species live widely in deep-sea and shallow-water areas, and thus grow piezophilically and piezosensitively. Piezophilic and psychrophilic Shewanella benthica cytochrome c 5 (SB cytc 5) was the most stable against guanidine hydrochloride (GdnHCl) and thermal denaturation, followed by less piezophilic but still psychrophilic Shewanella violacea cytochrome c 5 (SV cytc 5). These two were followed, as to stability level, by piezosensitive and mesophilic Shewanella amazonensis cytochrome c 5 (SA cytc 5), and piezosensitive and psychrophilic Shewanella livingstonensis cytochrome c 5 (SL cytc 5). The midpoint GdnHCl concentrations of SB cytc 5, SV cytc 5, SL cytc 5, and SA cytc 5 correlated with the optimal growth pressures of the species, the correlation coefficient value being 0.93. A similar trend was observed for thermal denaturation. Therefore, the stability of each cytochrome c 5 is related directly to its host's optimal growth pressure. Phylogenetic analysis indicated that Lys-37, Ala-41, and Leu-50 conserved in piezosensitive SL cytc 5 and SA cytc 5 are ancestors of the corresponding residues in piezophilic SB cytc 5 and SV cytc 5, Gln, Thr, and Lys, respectively, which might have been introduced during evolution on adaption to environmental pressure. The monomeric Shewanella cytochromes c 5 are suitable tools for examining protein stability with regard to the optimal growth pressures of the source species.
    Extremophiles 04/2014; · 2.20 Impact Factor
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    ABSTRACT: Sequence analysis indicated that thermophilic Hydrogenophilus thermoluteolus cytochrome c' (PHCP) and its mesophilic homolog, Allochromatium vinosum cytochrome c' (AVCP), closely resemble each other in a phylogenetic tree of the cytochrome c' family, with 55% sequence identity. The denaturation temperature of PHCP was 87 °C, 35 °C higher than that of AVCP. Furthermore, PHCP exhibited a larger enthalpy change value during its thermal denaturation than AVCP. While AVCP was dimeric, as observed previously, PHCP was trimeric, and this was the first observation as a cytochrome c'. Dissociation of trimeric PHCP and its protein denaturation reversibly occurred at the same time in a two-state transition manner. Therefore, PHCP is enthalpically more stable than AVCP, perhaps due to its unique trimeric form, in addition to the lower number of Gly residues in its putative α-helical regions.
    Bioscience Biotechnology and Biochemistry 08/2013; · 1.27 Impact Factor
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    ABSTRACT: Hydrogenophilus is a thermophilic, facultative chemoautotroph, which lives prevalently in high temperature geothermal niches. Despite the environmental distribution, little is known about its oxidative phosphorylation. Here, we show that inverted membrane vesicles derived from Hydrogenophilus thermoluteolus cells autotrophically cultivated with H2 formed a proton gradient on the addition of succinate, dl-lactate, and NADH, and exhibited oxidation activity toward these three organic compounds. These indicate the capability of mixotrophic growth of this bacterium. Biochemical analysis demonstrated that the same vesicles contained an F-type ATP synthase. The F1 sector of the ATP synthase purified from H. thermoluteolus membranes exhibited optimal ATPase activity at 65°C. Transformed Escherichia coli membranes expressing H. thermoluteolus F-type ATP synthase exhibited the same temperature optimum for the ATPase. These findings shed light on H. thermoluteolus oxidative phosphorylation from the aspects of membrane bioenergetics and ATPase biochemistry, which must be fundamental and advantageous in the biogeochemical cycles occurred in the high temperature geothermal niches.
    Environmental Microbiology Reports 04/2013; 5(2):235-42. · 3.26 Impact Factor
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    ABSTRACT: Cytochrome c₅ of pressure-sensitive Shewanella livingstonensis (SL cytc₅) exhibits lower thermal stability than a highly homologous counterpart of pressure-tolerant Shewanella violacea. This stability difference is due to an enthalpic effect that can be attributed to the amino acid residue at position 50 (Leu or Lys). These cytc₅ proteins are appropriate materials for understanding the protein stability mechanism.
    Bioscience Biotechnology and Biochemistry 09/2011; 75(9):1859-61. · 1.27 Impact Factor
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    ABSTRACT: Hyperthermophilic Aquifex aeolicus cytochrome c(555) (AA c(555)) exceptionally folds even in the apo state, unlike general cytochromes c including mesophilic Pseudomonas aeruginosa cytochrome c(551) (PA c(551)), which is structurally homologous to AA c(555) in the holo state. Here we hypothesized that the exceptional apo AA c(555) folding can be attributed to nine hydrophobic amino acid residues and proved this using a PA c(551) variant (denoted as PA-nh) carrying the nine hydrophobic residues at structurally corresponding positions. Circular dichroism experiments showed that the apo PA-nh variant became folded, unlike the wild-type apo PA c(551), and exhibited much higher stability than the wild type. Another difference between the holo forms of AA c(555) and PA c(551) is the existence of an extra helix in the former. Introduction of the amino acid residues forming the extra helix of AA c(555) into the PA-nh variant did not significantly affect its folding ability in the apo state. Therefore, the nine hydrophobic residues introduced into the apo PA-nh variant were enough to confer the folding ability. PA c(551) represents the first example of the conversion of an intrinsically unfolded apocytochrome c into an autonomously folded one, which was revealed by means of a protein engineering method without heme. Although heme is generally considered to be a trigger of apocytochrome c folding, the present results demonstrate a new heme-independent folding mechanism.
    Biochemistry 02/2011; 50(12):2313-20. · 3.38 Impact Factor