Characterization of hemin-binding protein 35 (HBP35) in Porphyromonas gingivalis: its cellular distribution, thioredoxin activity and role in heme utilization.
BACKGROUND
The periodontal pathogen Porphyromonas gingivalis is an obligate anaerobe that requires heme for growth. To understand its heme acquisition mechanism, we focused on a hemin-binding protein (HBP35 protein), possessing one thioredoxin-like motif and a conserved C-terminal domain, which are proposed to be involved in redox regulation and cell surface attachment, respectively.


RESULTS
We observed that the hbp35 gene was transcribed as a 1.1-kb mRNA with subsequent translation resulting in three proteins with molecular masses of 40, 29 and 27 kDa in the cytoplasm, and one modified form of the 40-kDa protein on the cell surface. A recombinant 40-kDa HBP35 exhibited thioredoxin activity in vitro and mutation of the two putative active site cysteine residues abolished this activity. Both recombinant 40- and 27-kDa proteins had the ability to bind hemin, and growth of an hbp35 deletion mutant was substantially retarded under hemin-depleted conditions compared with growth of the wild type under the same conditions.


CONCLUSIONS
P. gingivalis HBP35 exhibits thioredoxin and hemin-binding activities and is essential for growth in hemin-depleted conditions suggesting that the protein plays a significant role in hemin acquisition.
Shoji
Mikio
M
Division of Microbiology and Oral Infection, Department of Molecular Microbiology and Immunology, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan.
Shibata
Yasuko
Y
Shiroza
Teruaki
T
Yukitake
Hideharu
H
Peng
Benjamin
B
Chen
Yu-Yen
YY
Sato
Keiko
K
Naito
Mariko
M
Abiko
Yoshimitsu
Y
Reynolds
Eric C
EC
Nakayama
Koji
K
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D002352:Carrier Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D000081062:Heme-Binding Proteins; D006420:Hemeproteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D006427:Hemin / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D050505:Mutant Proteins / Q000378:metabolism; D016966:Porphyromonas gingivalis / Q000737:chemistry* / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D011485:Protein Binding; D014176:Protein Biosynthesis; D012329:RNA, Bacterial / Q000096:biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D013879:Thioredoxins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D002352:Carrier Proteins; D000081062:Heme-Binding Proteins; D006420:Hemeproteins; D050505:Mutant Proteins; D012329:RNA, Bacterial; D012333:RNA, Messenger; D013879:Thioredoxins; D006427:Hemin