The Extracellular Heme-binding Protein HbpS from the Soil Bacterium Streptomyces reticuli Is an Aquo-cobalamin Binder

Lucana, Darío Ortiz de Orué; Fedosov, Sergey N.; Wedderhoff, Ina; Edith, N.; Torda, Andrew E.

doi:10.1074/jbc.m114.585489

Cited by 9 publications

(12 citation statements)

References 68 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The K D -value of hemin binding by MBP-SpyB is lower than the reported affinities of heme-binding proteins involved in heme scavenging from host (Ortiz de Orué Lucana et al, 2014 ). However, the K D -value is within the range reported for some bacterial heme-sensing proteins, such as a regulator of porphyrin efflux system, PefR (Sachla et al, 2014 ), a repressor of photosystem genes, PpsR (Yin et al, 2012 ), light-regulated antirepressor, AppA (Yin et al, 2013 ), and a redox stress regulator, HbpS (Ortiz de Orué Lucana et al, 2014 ). Thus, these results suggest that SpyB plays a role in binding heme and possibly serves as an environmental sensor in GAS.…”

Section: Resultsmentioning

confidence: 60%

“…The rate of hemin dissociation from SpyB was determined using the procedure described in Ortiz de Orué Lucana et al ( 2014 ). Briefly, 5 μM MBP-SpyB was incubated with 20 μM hemin in 20 mM Tris-HCl, pH 7.5, 1 mM DTT at 4°C for 1 h. Free hemin was removed and MBP-SpyB:hemin holoprotein was collected by size-exclusion chromatography under the same conditions described above.…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

SpyB, a Small Heme-Binding Protein, Affects the Composition of the Cell Wall in Streptococcus pyogenes

Edgar

Chen

Kant

et al. 2016

Front. Cell. Infect. Microbiol.

View full text Add to dashboard Cite

Streptococcus pyogenes (Group A Streptococcus or GAS) is a hemolytic human pathogen associated with a wide variety of infections ranging from minor skin and throat infections to life-threatening invasive diseases. The cell wall of GAS consists of peptidoglycan sacculus decorated with a carbohydrate comprising a polyrhamnose backbone with immunodominant N-acetylglucosamine side-chains. All GAS genomes contain the spyBA operon, which encodes a 35-amino-acid membrane protein SpyB, and a membrane-bound C3-like ADP-ribosyltransferase SpyA. In this study, we addressed the function of SpyB in GAS. Phenotypic analysis of a spyB deletion mutant revealed increased bacterial aggregation, and reduced sensitivity to β-lactams of the cephalosporin class and peptidoglycan hydrolase PlyC. Glycosyl composition analysis of cell wall isolated from the spyB mutant suggested an altered carbohydrate structure compared with the wild-type strain. Furthermore, we found that SpyB associates with heme and protoporphyrin IX. Heme binding induces SpyB dimerization, which involves disulfide bond formation between the subunits. Thus, our data suggest the possibility that SpyB activity is regulated by heme.

show abstract

Section: Resultsmentioning

confidence: 60%

Section: Methodsmentioning

confidence: 99%

SpyB, a Small Heme-Binding Protein, Affects the Composition of the Cell Wall in Streptococcus pyogenes

Edgar

Chen

Kant

et al. 2016

Front. Cell. Infect. Microbiol.

View full text Add to dashboard Cite

show abstract

“…PduOC (10 μM) was incubated with 20 μM of either aquo-cobalamin (H 2 OCbl), methylcobalamin (MeCbl), (AdoCbl) or cyanocobalamin (CNCbl) at 37°C for 2 h. Mixtures were loaded on to a native polyacrylamide (PAA) gel that was photographed after ~30 min of electrophoresis and scanned inmediately after electrophoresis. Binding was also assessed by mixing H 2 OCbl (10 μM) with 20 μM of the apoprotein and recording UV/Vis spectra after 2 h (Ortiz de Orué Lucana et al, 2014 ). The H 2 OCbl-binding protein, HpbS (Ortiz de Orué Lucana et al, 2014 ) was used to check conditions as a positive control.…”

Section: Methodsmentioning

confidence: 99%

“…Binding was also assessed by mixing H 2 OCbl (10 μM) with 20 μM of the apoprotein and recording UV/Vis spectra after 2 h (Ortiz de Orué Lucana et al, 2014 ). The H 2 OCbl-binding protein, HpbS (Ortiz de Orué Lucana et al, 2014 ) was used to check conditions as a positive control.…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

The Crystal Structure of the C-Terminal Domain of the Salmonella enterica PduO Protein: An Old Fold with a New Heme-Binding Mode

et al. 2016

Self Cite

View full text Add to dashboard Cite

The two-domain protein PduO, involved in 1,2-propanediol utilization in the pathogenic Gram-negative bacterium Salmonella enterica is an ATP:Cob(I)alamin adenosyltransferase, but this is a function of the N-terminal domain alone. The role of its C-terminal domain (PduOC) is, however, unknown. In this study, comparative growth assays with a set of Salmonella mutant strains showed that this domain is necessary for effective in vivo catabolism of 1,2-propanediol. It was also shown that isolated, recombinantly-expressed PduOC binds heme in vivo. The structure of PduOC co-crystallized with heme was solved (1.9 Å resolution) showing an octameric assembly with four heme moieities. The four heme groups are highly solvent-exposed and the heme iron is hexa-coordinated with bis-His ligation by histidines from different monomers. Static light scattering confirmed the octameric assembly in solution, but a mutation of the heme-coordinating histidine caused dissociation into dimers. Isothermal titration calorimetry using the PduOC apoprotein showed strong heme binding (Kd = 1.6 × 10−7 M). Biochemical experiments showed that the absence of the C-terminal domain in PduO did not affect adenosyltransferase activity in vitro. The evidence suggests that PduOC:heme plays an important role in the set of cobalamin transformations required for effective catabolism of 1,2-propanediol. Salmonella PduO is one of the rare proteins which binds the redox-active metabolites heme and cobalamin, and the heme-binding mode of the C-terminal domain differs from that in other members of this protein family.

show abstract

Characterization of the complex between native and reduced bovine serum albumin with aquacobalamin and evidence of dual tetrapyrrole binding

et al. 2018

View full text Add to dashboard Cite

Serum albumin binds to a variety of endogenous ligands and drugs. Human serum albumin (HSA) binds to heme via hydrophobic interactions and axial coordination of the iron center by protein residue Tyr161. Human serum albumin binds to another tetrapyrrole, cobalamin (Cbl), but the structural and functional properties of this complex are poorly understood. Herein, we investigate the reaction between aquacobalamin (HOCbl) and bovine serum albumin (BSA, the bovine counterpart of HSA) using Ultraviolet-Visible and fluorescent spectroscopy, and electron paramagnetic resonance. The reaction between HOCbl and BSA led to the formation of a BSA-Cbl(III) complex consistent with N-axial ligation (amino). Prior to the formation of this complex, the reactants participate in an additional binding event that has been examined by fluorescence spectroscopy. Binding of BSA to Cbl(III) reduced complex formation between the bound cobalamin and free cyanide to form cyanocobalamin (CNCbl), suggesting that the β-axial position of the cobalamin may be occupied by an amino acid residue from the protein. Reaction of BSA containing reduced disulfide bonds with HOCbl produces cob(II)alamin and disulfide with intermediate formation of thiolate Cbl(III)-BSA complex and its decomposition. Finally, in vitro studies showed that cobalamin binds to BSA only in the presence of an excess of protein, which is in contrast to heme binding to BSA that involves a 1:1 stoichiometry. In vitro formation of BSA-Cbl(III) complex does not preclude subsequent heme binding, which occurs without displacement of HOCbl bound to BSA. These data suggest that the two tetrapyrroles interact with BSA in different binding pockets.

show abstract

The Extracellular Heme-binding Protein HbpS from the Soil Bacterium Streptomyces reticuli Is an Aquo-cobalamin Binder

Cited by 9 publications

References 68 publications

SpyB, a Small Heme-Binding Protein, Affects the Composition of the Cell Wall in Streptococcus pyogenes

SpyB, a Small Heme-Binding Protein, Affects the Composition of the Cell Wall in Streptococcus pyogenes

The Crystal Structure of the C-Terminal Domain of the Salmonella enterica PduO Protein: An Old Fold with a New Heme-Binding Mode

Characterization of the complex between native and reduced bovine serum albumin with aquacobalamin and evidence of dual tetrapyrrole binding

Contact Info

Product

Resources

About