Sulfide-Binding Hemoglobins: Effects of Mutations on Active-Site Flexibility

Fernández-Alberti, Sebastián; Bacelo, Daniel E.; Binning, R. C.; Echave, Julián; Chergui, Majed; López‐Garriga, Juan

doi:10.1529/biophysj.106.081646

Cited by 21 publications

(23 citation statements)

References 54 publications

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“…These association rate constants are also comparable with those reported for the wt-HbI, k on of 2.3 × 10 5 /M/s [17]. As previously pointed out by Fernández-Alberti, et al [39]. , the fast sulfide association rate constant is highly affected by changes in the HbI distal pocket and its periphery.…”

Section: Resultssupporting

confidence: 89%

Molecular Cloning and Characterization of a (Lys)6-Tagged Sulfide-Reactive Hemoglobin I from Lucina pectinata

et al. 2015

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A poly-Lys tag was fused to the Lucina pectinata hemoglobin I (HbI) coding sequence and purified using an efficient and fast process. HbI is a hemeprotein that binds hydrogen sulfide (H2S) with high affinity and it has been used to understand physiologically relevant reactions of this signaling molecule. The (Lys)6-tagged rHbI construct was expressed in E. coli and purified by immobilization on a cation exchange matrix, followed by size-exclusion chromatography. The identity, structure, and function of the (Lys)6-tagged rHbI were assessed by mass spectrometry, small and wide X-ray scattering, optical spectroscopy, and kinetic analysis. The scattering and spectroscopic results showed that the (Lys)6-tagged rHbI is structurally and functionally analogous to the native protein as well as to the (His)6-tagged rHbI. Kinetics studies with H2S indicated that the association (kon) and dissociation (koff) rate constants were 1.4 × 105/M/s and 0.1 × 10−3/s, respectively. This results confirmed that the (Lys)6-tagged rHbI binds H2S with the same high affinity as its homologue.

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Section: Resultssupporting

confidence: 89%

Molecular Cloning and Characterization of a (Lys)6-Tagged Sulfide-Reactive Hemoglobin I from Lucina pectinata

et al. 2015

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“…Moreover, among the three Hbs present in Lucina pectinata (I, II, and III), only HbI shows a very high sulfide affinity, and it has been proposed that the presence of TyrB10 in HbII and HbIII (PheB10 in HbI) destabilizes the binding of sulfide. 63 These results depict the complexity of the distal-side scenario that is under active discussion. As explained …”

Section: ■ Discussionmentioning

confidence: 93%

Reactivity of Inorganic Sulfide Species toward a Heme Protein Model

et al. 2014

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The reactivity of inorganic sulfide species toward heme peptides was explored under biorelevant conditions in order to unravel the molecular details of the reactivity of the endogenous hydrogen sulfide toward heme proteins. Unlike ferric porphyrinates, which are reduced by inorganic sulfide, some heme proteins can form stable Fe(III)-sulfide adducts. To isolate the protein factors ruling the redox chemistry, we used as a system model, the undecapeptide microperoxidase (MP11), a heme peptide derived from cytochrome c proteolysis that retains the proximal histidine bound to the Fe(III) atom. Upon addition of gaseous hydrogen sulfide (H2S) at pH 6.8, the UV-vis spectra of MP11 closely resembled those of the low-spin ferric hydroxo complex (only attained at an alkaline pH) and cysteine or alkylthiol derivatives, suggesting that the Fe(III) reduction was prevented. The low-frequency region of the resonance Raman spectrum revealed the presence of an Fe(III)-S band at 366 cm(-1) and the general features of a low-spin hexacoordinated heme. Anhydrous sodium sulfide (Na2S) was the source of sulfide of choice for the kinetic evaluation of the process. Theoretical calculations showed no distal stabilization mechanisms for bound sulfide species in MP11, highlighting a key role of the proximal histidine for the stabilization of the Fe(III)-S adducts of heme compounds devoid of distal counterparts, which is significant with regard to the biochemical reactivity of endogenous hydrogen sulfide.

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“…Similarly, a large cavity volume was observed in the ferric derivative of the Mb triple mutant (L29F/H64Q/V68F), Mb, and HbI LP . The crystal structure reveals the existence of a much greater heme freedom and larger distal cavity volume in HbI LP than sperm whale Mb, in both the H 2 S bound and unbound to the heme group, because of the lack of hydrogen bonding between the heme propionate groups and nearby amino acid residues (43). This dynamic behavior is absent in HbII LP , where the heme group is firmly anchored in place.…”

Section: Discussionmentioning

confidence: 93%