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HmuY haemophore and gingipain proteases constitute a unique syntrophic system of haem acquisition by Porphyromonas gingivalis.

Smalley JW, Byrne DP, Birss AJ, Wojtowicz H, Sroka A, Potempa J, Olczak T - PLoS ONE (2011)

Bottom Line: HmuY was also capable of scavenging haem from oxyhaemoglobin pre-treated with the K-gingipain (Kgp).This is the first demonstration of a haemophore working in conjunction with proteases to acquire haem from haemoglobin.In addition, HmuY was able to extract haem from methaemalbumin, and could bind haem, either free in solution or from methaemoglobin, even in the presence of serum albumin.

View Article: PubMed Central - PubMed

Affiliation: Department of Clinical Infection, Microbiology and Immunology, Institute of Infection and Global Health, [corrected] University of Liverpool, Liverpool, United Kingdom. josmall@liv.ac.uk

ABSTRACT
Haem (iron protoporphyrin IX) is both an essential growth factor and virulence regulator for the periodontal pathogen Porphyromonas gingivalis, which acquires it mainly from haemoglobin via the sequential actions of the R- and K-specific gingipain proteases. The haem-binding lipoprotein haemophore HmuY and its cognate receptor HmuR of P. gingivalis, are responsible for capture and internalisation of haem. This study examined the role of the HmuY in acquisition of haem from haemoglobin and the cooperation between HmuY and gingipain proteases in this process. Using UV-visible spectroscopy and polyacrylamide gel electrophoresis, HmuY was demonstrated to wrest haem from immobilised methaemoglobin and deoxyhaemoglobin. Haem extraction from oxyhaemoglobin was facilitated after oxidation to methaemoglobin by pre-treatment with the P. gingivalis R-gingipain A (HRgpA). HmuY was also capable of scavenging haem from oxyhaemoglobin pre-treated with the K-gingipain (Kgp). This is the first demonstration of a haemophore working in conjunction with proteases to acquire haem from haemoglobin. In addition, HmuY was able to extract haem from methaemalbumin, and could bind haem, either free in solution or from methaemoglobin, even in the presence of serum albumin.

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Related in: MedlinePlus

Formation of the ferrohaem-HmuY complex during incubation of HmuY with deoxyhaemoglobin-agarose.Haemoglobin-agarose was deoxygenated with sodium dithionite and maintained anaerobically during the reaction with HmuY. Concentration of haemoglobin was 16 µM (with respect to haemoglobin subunit) as was HmuY. See text for details. The absorbance below 375 nm is due to the presence of dithionite. A small amount of deoxyhaemoglobin (∼0.1% of the total) was released from the control deoxyhaemoglobin-agarose after 40 min incubation in the absence of HmuY.
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pone-0017182-g010: Formation of the ferrohaem-HmuY complex during incubation of HmuY with deoxyhaemoglobin-agarose.Haemoglobin-agarose was deoxygenated with sodium dithionite and maintained anaerobically during the reaction with HmuY. Concentration of haemoglobin was 16 µM (with respect to haemoglobin subunit) as was HmuY. See text for details. The absorbance below 375 nm is due to the presence of dithionite. A small amount of deoxyhaemoglobin (∼0.1% of the total) was released from the control deoxyhaemoglobin-agarose after 40 min incubation in the absence of HmuY.

Mentions: Since P. gingivalis may also encounter deoxygenated haemoglobin in the anaerobic gingival sulcus or periodontal pocket, it was decided to investigate the interaction of HmuY with deoxyhaemoglobin-agarose. The haemoglobin-agarose was deoxygenated by the addition of sodium dithionite [19] and maintained anaerobically whilst periodically removing the agarose-free, HmuY-containing supernatants for which the spectra were recorded. These showed that a ferrohaem-HmuY complex developed within 5 min of exposure of the deoxyhaemoglobin to HmuY (Fig. 10).


HmuY haemophore and gingipain proteases constitute a unique syntrophic system of haem acquisition by Porphyromonas gingivalis.

Smalley JW, Byrne DP, Birss AJ, Wojtowicz H, Sroka A, Potempa J, Olczak T - PLoS ONE (2011)

Formation of the ferrohaem-HmuY complex during incubation of HmuY with deoxyhaemoglobin-agarose.Haemoglobin-agarose was deoxygenated with sodium dithionite and maintained anaerobically during the reaction with HmuY. Concentration of haemoglobin was 16 µM (with respect to haemoglobin subunit) as was HmuY. See text for details. The absorbance below 375 nm is due to the presence of dithionite. A small amount of deoxyhaemoglobin (∼0.1% of the total) was released from the control deoxyhaemoglobin-agarose after 40 min incubation in the absence of HmuY.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3040768&req=5

pone-0017182-g010: Formation of the ferrohaem-HmuY complex during incubation of HmuY with deoxyhaemoglobin-agarose.Haemoglobin-agarose was deoxygenated with sodium dithionite and maintained anaerobically during the reaction with HmuY. Concentration of haemoglobin was 16 µM (with respect to haemoglobin subunit) as was HmuY. See text for details. The absorbance below 375 nm is due to the presence of dithionite. A small amount of deoxyhaemoglobin (∼0.1% of the total) was released from the control deoxyhaemoglobin-agarose after 40 min incubation in the absence of HmuY.
Mentions: Since P. gingivalis may also encounter deoxygenated haemoglobin in the anaerobic gingival sulcus or periodontal pocket, it was decided to investigate the interaction of HmuY with deoxyhaemoglobin-agarose. The haemoglobin-agarose was deoxygenated by the addition of sodium dithionite [19] and maintained anaerobically whilst periodically removing the agarose-free, HmuY-containing supernatants for which the spectra were recorded. These showed that a ferrohaem-HmuY complex developed within 5 min of exposure of the deoxyhaemoglobin to HmuY (Fig. 10).

Bottom Line: HmuY was also capable of scavenging haem from oxyhaemoglobin pre-treated with the K-gingipain (Kgp).This is the first demonstration of a haemophore working in conjunction with proteases to acquire haem from haemoglobin.In addition, HmuY was able to extract haem from methaemalbumin, and could bind haem, either free in solution or from methaemoglobin, even in the presence of serum albumin.

View Article: PubMed Central - PubMed

Affiliation: Department of Clinical Infection, Microbiology and Immunology, Institute of Infection and Global Health, [corrected] University of Liverpool, Liverpool, United Kingdom. josmall@liv.ac.uk

ABSTRACT
Haem (iron protoporphyrin IX) is both an essential growth factor and virulence regulator for the periodontal pathogen Porphyromonas gingivalis, which acquires it mainly from haemoglobin via the sequential actions of the R- and K-specific gingipain proteases. The haem-binding lipoprotein haemophore HmuY and its cognate receptor HmuR of P. gingivalis, are responsible for capture and internalisation of haem. This study examined the role of the HmuY in acquisition of haem from haemoglobin and the cooperation between HmuY and gingipain proteases in this process. Using UV-visible spectroscopy and polyacrylamide gel electrophoresis, HmuY was demonstrated to wrest haem from immobilised methaemoglobin and deoxyhaemoglobin. Haem extraction from oxyhaemoglobin was facilitated after oxidation to methaemoglobin by pre-treatment with the P. gingivalis R-gingipain A (HRgpA). HmuY was also capable of scavenging haem from oxyhaemoglobin pre-treated with the K-gingipain (Kgp). This is the first demonstration of a haemophore working in conjunction with proteases to acquire haem from haemoglobin. In addition, HmuY was able to extract haem from methaemalbumin, and could bind haem, either free in solution or from methaemoglobin, even in the presence of serum albumin.

Show MeSH
Related in: MedlinePlus