Site-Directed Mutagenesis of Bacterial Hemoglobin: The Role of Glutamine (E7) in Oxygen-Binding in the Distal Heme Pocket

Dikshit, Kanak L.; Orii, Yutaka; Navani, Naveen Kumar; Patel, S. M.; Huang, Hui‐Yu; Stark, Benjamin C.; Webster, D. R.

doi:10.1006/abbi.1997.0432

Cited by 42 publications

(35 citation statements)

References 29 publications

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“…A similar stabilization of O 2 by Tyr(B10) is seen in Ascaris Hb [164][165][166], M. tuberculosis HbN and other Hbs [167][168][169][170] suggesting similar adaptation for a NOD function. In addition, highly conserved Gln(E7) in the flexible CD loop and more distant from heme in flavoHbs and Vitreoscilla Hb may form a hydrogen bond network stabilizing O 2 binding [157,158,171,172], although evidence for O 2 binding by E7 mutants of Vitreoscilla Hb suggests otherwise [172]. A similar role has been described for Gln(E7) in Ascaris Hb and other Hbs [164][165][166].…”

Section: Flavohb Hb and Mb Structures And The No Dioxygenase Mechanismmentioning

confidence: 70%

Nitric oxide dioxygenase function and mechanism of flavohemoglobin, hemoglobin, myoglobin and their associated reductases

Gardner

2005

Journal of Inorganic Biochemistry

244

187

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Section: Flavohb Hb and Mb Structures And The No Dioxygenase Mechanismmentioning

confidence: 70%

Nitric oxide dioxygenase function and mechanism of flavohemoglobin, hemoglobin, myoglobin and their associated reductases

Gardner

2005

Journal of Inorganic Biochemistry

244

187

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“…As a result, the E7 Gln is rotated out of the heme pocket. Spectral and kinetic studies of the binding of oxygen and CO to E7 mutants of Vitreoscilla hemoglobin showed that this substitution had little effect on the ligand binding properties of this protein (18), evidence that E7 Gln does not hydrogen-bond to the bound ligand. The same segment in another bacterium, Alcaligenes eutrophus, adopts an elongated and well defined E-helix but deviates significantly from that in other globins by more than 10°because of an increase in the angle between E and F helices (19).…”

Section: Discussionmentioning

confidence: 98%

A Cooperative Oxygen Binding Hemoglobin from Mycobacterium tuberculosis

Yeh¹,

Couture²,

Ouellet³

et al. 2000

Journal of Biological Chemistry

120

139

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The homodimeric hemoglobin (HbN) from Mycobacterium tuberculosis displays an extremely high oxygen binding affinity and cooperativity. Sequence alignment with other hemoglobins suggests that the proximal F8 ligand is histidine, the distal E7 residue is leucine, and the B10 position is occupied by tyrosine. To determine how these heme pocket residues regulate the ligand binding affinities and physiological functions of HbN, we have measured the resonance Raman spectra of the O 2 , CO, and OH ؊ derivatives of the wild type protein and the B10 Tyr 3 Leu and Phe mutants. Taken together these data demonstrate a unique distal environment in which the heme bound ligands strongly interact with the B10 tyrosine residue. The implications of these data on the physiological functions of HbN and another heme-containing protein, cytochrome c oxidase, are considered.

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“…Subcloning of vgb into pEG202 and cyo Genes and Gene Fragments into pJG4-5-vgb was amplified by PCR from pNKD1 (15), which bears wild type vgb, using the primers 5Ј-GCGCGGAATTCATGTTAGACCA-GCAAACCATTAACATCATC-3Ј and 5Ј-GCGCGCTCGAGTTATTCAA-CCGCTTGAGCGTACAAATCTG-3Ј. After restriction digestion of the PCR fragment with EcoRI and XhoI, vgb was inserted into pEG202.…”

Section: Methodsmentioning

confidence: 99%

Vitreoscilla Hemoglobin Binds to Subunit I of Cytochrome bo Ubiquinol Oxidases

Park

Kim

Howard

et al. 2002

Journal of Biological Chemistry

Self Cite

106

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The bacterium, Vitreoscilla, can induce the synthesis of a homodimeric hemoglobin under hypoxic conditions. Expression of VHb in heterologous bacteria often enhances growth and increases yields of recombinant proteins and production of antibiotics, especially under oxygen-limiting conditions. There is evidence that VHb interacts with bacterial respiratory membranes and cytochrome bo proteoliposomes. We have examined whether there are binding sites for VHb on the cytochrome, using the yeast two-hybrid system with VHb as the bait and testing every Vitreoscilla cytochrome bo subunit as well as the soluble domains of subunits I and II. A significant interaction was observed only between VHb and intact subunit I. We further examined whether there are binding sites for VHb on cytochrome bo from Escherichia coli and Pseudomonas aeruginosa, two organisms in which stimulatory effects of VHb have been observed. Again, in both cases a significant interaction was observed only between VHb and subunit I. Because subunit I contains the binuclear center where oxygen is reduced to water, these data support the function proposed for VHb of providing oxygen directly to the terminal oxidase; it may also explain its positive effects in Vitreoscilla as well as in heterologous organisms.

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Site-Directed Mutagenesis of Bacterial Hemoglobin: The Role of Glutamine (E7) in Oxygen-Binding in the Distal Heme Pocket

Cited by 42 publications

References 29 publications

Nitric oxide dioxygenase function and mechanism of flavohemoglobin, hemoglobin, myoglobin and their associated reductases

Nitric oxide dioxygenase function and mechanism of flavohemoglobin, hemoglobin, myoglobin and their associated reductases

A Cooperative Oxygen Binding Hemoglobin from Mycobacterium tuberculosis

Vitreoscilla Hemoglobin Binds to Subunit I of Cytochrome bo Ubiquinol Oxidases

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