• BMB Reports
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• 제45권4호
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• pp.239-241
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• 2012

Iron availability is limited in the environment and most bacteria have developed a system to acquire iron from host hemoproteins. Heme oxygenase plays an important role by degrading heme group and releasing the essential nutrient iron. The structure of Bacillus subtilis HmoB was determined to 2.0 ${\AA}$ resolution. B. subtilis HmoB contains a typical antibiotic biosynthesis monooxygenase (ABM) domain that spans from 71 to 146 residues and belongs to the IsdG family heme oxygenases. Comparison of HmoB and IsdG family proteins showed that the C-terminal region of HmoB has similar sequence and structure to IsdG family proteins and contains conserved critical residues for heme degradation. However, HmoB is distinct from other IsdG family proteins in that HmoB is about 60 amino acids longer in the N-terminus and does not form a dimer whereas previously studied IsdG family heme oxygenases form functional homodimers. Interestingly, the structure of monomeric HmoB resembles the dimeric structure of IsdG family proteins. Hence, B. subtilis HmoB is a heme oxygenase with a novel structural feature.

• Bulletin of the Korean Chemical Society
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• 제27권11호
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• pp.1825-1831
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• 2006

The influence of solvent viscosity on conformational dynamics of the heme-pocket, a small vacant site near the binding site of myoglobin (Mb) and hemoglobin (Hb), and playing a functionally important role by serving as a station in ligand binding and escape, was studied by probing time-resolved vibrational spectra of CO photodissociated from MbCO and HbCO in $D_2O$, 75 wt% glycerol/$D_2O$, and trehalose at 283 K. Two absorption bands ($B_1$ and $B_2$) of the sample in viscous solvents, arising from CO in the heme pocket, are very similar to those in $D_2O$. Two bands in Mb and Hb under all three solvents exhibit very similar nonexponential spectral evolution ($B_1$ band; blue shifting and broadening, $B_2$ band; narrowing with a negligible shifting), suggesting that in the present experimental time window of 100 ps, the extents of the spectral shift and narrowing is much influenced neither by the viscosity of solvent nor by the quaternary contact of Hb. Spectral evolution can be described by a biexponential function with a fast universal time constant of 0.52 ps and a slow time constant ranging from 13 to 32 ps. For both proteins in all three solvents majority of spectral evolution occurs with the fast universal time constant. The magnitude of the slow rate in the spectral shift of B1 band decreases with increasing solvent viscosity, indicating that it is influenced by global conformational change which is retarded in viscous solvent, thereby serve as a reporter of global conformational change of heme proteins after deligation.

• BMB Reports
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• 제31권4호
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• pp.307-327
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• 1998

Cytochrome c peroxidase (CcP) is a yeast mitochondrial enzyme which catalyzes the reduction of hydrogen peroxide to water using two equivalents of ferrocytochrome c. The CcP/cytochrome c system has many features which make it a very useful model for detailed investigation of heme protein structure/function relationships including activation of hydrogen peroxide, protein-protein interactions, and long-range electron transfer. Both CcP and cytochrome c are single heme, single subunit proteins of modest size. High-resolution crystallographic structures of both proteins, of one-to-one complexes of the two proteins, and a number of active-site mutants are available. Site-directed mutagenesis studies indicate that the distal histidine in CcP is primarily responsible for rapid utilization of hydrogen peroxide implying significantly different properties of the distal histidine in the peroxidases compared to the globins. CcP and cytochrome c bind to form a dynamic one-to-one complex. The binding is largely electrostatic in nature with a small, unfavorable enthalpy of binding and a large positive entropy change upon complex formation. The cytochrome c-binding site on CcP has been mapped in solution by measuring the binding affinities between cytochrome c and a number of CcP surface mutations. The binding site for cytochrome c in solution is consistent with the crystallographic structure of the one-to-one complex. Evidence for the involvement of a second, low-affinity cytochrome c-binding site on CcP in long-range electron transfer between the two proteins is reviewed.

• Bulletin of the Korean Chemical Society
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• 제26권1호
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• pp.151-156
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• 2005

The conformational dynamics of heme pocket, a small vacant site near the binding site of heme proteins -myoglobin (Mb) and hemoglobin (Hb), was investigated after photolysis of carbon monoxide from MbCO and HbCO in D$_2$O solution at 283 K by probing time-resolved vibrational spectra of photolyzed CO. Two absorption bands, arising from CO in the heme pocket, evolve nonexponentially in time. The band at higher energy side blue shifts and broadens with time and the one at lower energy side narrows significantly with a negligible shift. These spectral evolutions are induced by protein conformational changes following photolysis that modify structure and electric field of heme pocket, and ligand dynamics in it. The conformational changes affecting the spectrum of photolyzed CO in heme pocket likely modulates ligand-binding activity.

• Bulletin of the Korean Chemical Society
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• 제23권6호
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• pp.865-872
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• 2002

A systematic way of determining the equilibrium orientation of carbon monoxide (CO) in heme proteins using time-resolved polarized mid-IR spectroscopy is presented. The polarization anisotropy at pump-probe delay time of zero in the limit of zero photolysis and the angular distrbution function of CO are required to obtain the equilibrium orientation of CO. An approach is developed for determining the polarization anisotropy in the zero-photolysis limit from the anisotropy measured under finite photolysis conditions in an optically thick sample where the fraction of molecules photolyzed decreased as the pump pulse passes through and is absorbed by the sample. This approach is verified by measuring the polarization anisotropy of CO of carbonmonoxy myoglobin at various levels of photolysis. This method can be readily applied to other photoselection experiments determining precise angle between transition dipoles.

• Journal of Nutrition and Health
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• 제33권2호
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• pp.167-178
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• 2000

The effect of oral vitamin e (800IU/day) and C (500mg/day) supplementation for 10 days and/or smoking cessation for 5 days on oxidative damage to the red blood cells (RBC) of male smokers (22.2$\pm$0.2 years old) was studied. RBC were tested for their ability to protect against smoking-induced oxidative damage by measuring heme proteins (carboxyhemoglobin, hemoglobin, methemoglobin, oxyhemoglobin), hemolysis and thiobarbiturinc acid reactive substances (TBARS). Plasma levels of vitamin c, A, E, $\beta$-catotene, total cholesterol, glutamic pyruvic transaminase(GPT) and glutamic oxaloacetic transaminase(GOT) were also analyzed. In experiment one, a comparison was made of heme proteins and lipid damage to RBC, plasma antioxidant status (indexed by plasma levels of vitamin C, E, A and $\beta$-carotene) between smokers(n=56) and non-smokers (n=16). No differences were found in plasma antioxidant status, heme protein damage and TBARS concentration of RBC. In experiment two, 46 fasting male smokers from experiment one were divided into 4 groups. The groups were smoking with placebo group(SP, n=14), smoking cessation with vitamins supplementatin group (SV, n=13), smoking cessation with placebo group (NSP, n=9) and smoking cessation with vitamins supplementation group (NSV, n=10). After supplementing antioxidant vitamins, significant increases were seen in plasma vitamins supplementation group (NSV, n=10). After supplementing antioxidant vitamins, significant increases were seen plasma vitamin C (p<0.05) and vitamin E levels (p<0.05). The plasma vitamin E level was highest in the NSV group. Vitmain E and C supplementation provided some protection against heme proteins and lipid damage by lowering methemoglobin, hemolysis and TBARS concentration of RBC. Smoking cessation significantly decreased TBARS of RBC and plasma total cholesterol concentration. Supplementing vitamin E and C with smoking cessation considerably lowered plasma total cholesterol. These results point to a special association among smoking, oxidative damage and plasma antioxidant vitamin status. They indicate that increases in plasma antioxidant status can be detected after the supplementation of vitamin C and E and that smoking cessation had an additional effect on plasma vitamin E level. The present data suggest that improved antioxidant status induced by antioxidant supplementation or smoking cessation may help prevent oxidative damage in smokers.

• Archives of Pharmacal Research
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• 제29권3호
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• pp.249-255
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• 2006

Ability of iron acquisition of pathogenic microorganisms functions as a virulence factor. Candida albicans, a fungal pathogen that requires iron for growth, is susceptible to growth retardation by high-affinity iron binding proteins such as transferrin. Recently, we reported that C. albicans could utilize the heme as a part of heme-containing proteins dissociated by heme oxygenase, CaHMX1. In search of another pathway that C. albicans can use to bypass the growth regulation produced by iron limitation, this present study examined utilization of non-candidal siderophores such as Desferal and rhodotorulic acid (RA) for acquisition of inorganic iron by the fungus. C. albicans secreting no siderophores was cultured in iron-free (pretreated with apotransferrin for 24 h) (culture medium). Once growth of the yeast reached stasis from iron starvation, a siderophore was added to the culture media. Results showed that cultures containing apotransferrin within a dialysis membrane recovered growth to the level of untreated controls, whereas C. albicans yeast cells in direct contact with soluble iron-free (apo) transferrin recovered growth only partially. When static growth from iron limitation was reached, the addition of siderophore-apotransferrin complex to culture medium also permitted the yeast to recover growth from apotransferrin growth regulation. All the data show that C. albicans can utilize the non-candidal siderophores for iron acquisition under transferrin regulation as can pathogenic bacteria.

• Journal of Plant Biology
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• 제36권3호
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• pp.301-304
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• 1993

Root nodule specific proteins of Alnus hirsuta were examined. SDS-PAGE pattern of the Alnus root nodule was simpler than that of soybean, showing five nodule specific proteins whose molecular weights were 48, 40, 36, 26 and 19 kD, respectively. Among them, 48 kD protein existed most abundantly and were composed of two subunits whose pI value were 4.0 and 4.3, respectively. The 48 kD protein seemed to be a heme containing protein based on reaction with diaminobenzidine. Although 19 kD protein was present in small amount, it was most similar to leghemoglobin in terms of its molecular weight.

• Archives of Pharmacal Research
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• 제29권1호
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• pp.50-58
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• 2006

Translationally controlled tumor protein (TCTP), also known as histamine releasing factor (HRF), is found abundantly in different eukaryotic cell types. The sequence homology of TCTP between different species is very high, belonging to the MSS4/DSS4 superfamily of proteins. TCTP is involved in both cell growth and human late allergy reaction, as well as having a calcium binding property; however, its primary biological functions remain to be clearly elucidated. In regard to many possible functions, the TCTP of Plasmodium falciparum (Pf) is known to bind with an antimalarial agent, artemisinin, which is activated by heme. It is assumed that the endoperoxide-bridge of artemisinin is opened up by heme to form a free radical, which then eventually alkylates, probably to the Cys14 of PfTCTP. Study of the docking of artemisinin with heme, and subsequently with PfTCTP, was carried out to verify the above hypothesis on the basis of structural interactions. The three dimensional (3D) structure of PfTCTP was built by homology modeling, using the NMR structure of the TCTP of Schizosaccharomyces pombe as a template. The quality of the model was examined based on its secondary structure and biological function, as well as with the use of structure evaluating programs. The interactions between artemisinin, heme and PfTCTP were then studied using the docking program, FlexiDock. The center of the peroxide bond of artemisinin and the Fe of heme were docked within a short distance of $2.6{\AA}$, implying the strong possibility of an interaction between the two molecules, as proposed. When the activated form of artemisinin was docked on the PfTCTP, the C4-radical of the drug faced towards the sulfur of Cys14 within a distance of $2.48{\AA}$, again suggesting the possibility of alkylation having occurred. These results confirm the proposed mechanism of the antimalarial effect of artemisinin, which will provide a reliable method for establishing the mechanism of its biological activity using a molecular modeling study.

• BMB Reports
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• 제34권4호
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• pp.385-389
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• 2001

Insulin-like growth factor binding protein-1 (IGFBP-1) appears to be an important modular of the insulin growth factor (IGF) bioactivity in metabolic disease and chronic hypoxia. Treatment of desferrioxamine (Dfo), cobalt, or nickel in HepG2 cells stimulated the expression of IGFBP1 mRNA as hypoxia. However, the presence of ferric ammonium citrate (FAC) in the 1% $O_2$ decreased the upregulation of the IGFBP-1 mRNA expression. In addition, actinomycin D and cycloheximide abolished the increase in the expression of IGFBP-1 mRNA that was induced by Dfo and transition metals (cobalt and nickel). To obtain further information about the putative oxygen sensor, we postulate that putative heme proteins, responsible for the oxygen-sensing process in HepG2 cells, should be sensitive to hypoada. The mechanism of these upregulations of the IGFBP-1 mRNA expression by Dfo and transition metals was investigated by treatment with 2 mM of 4,6-dioxoheptanoic acid (DHA), an inhibitor of heme biosynthesis. The results showed that 1% $O_2$-, Dfo-, cobalt-, or nickel induced IGFBP-1 mRNA expressions in HepG2 cells were all markedly inhibited when the heme synthesis was blocked by DHA. We suggest that the IGFBP-1 mRNA expression in the HepG2 cell is regulated by 1% $O_2$, Dfo, cobalt, or nickel, implicating the involvement of the putative heme-containing oxygensensing molecule.