Porphyromonas gingivalis, an oral bacterium associated with periodontal disease, requires haemin for growth. Although several multigenic clusters encoding haemin-uptake systems are present on the genome of P. gingivalis, little is known regarding their transcriptional organization and expression. This study identified a 23 kDa iron-regulated haemin-binding protein encoded by a larger than previously reported variant of hmuY. It was shown that the hmu locus is larger than previously reported and is composed of six genes, hmuYRSTUV, encoding a novel hybrid haemin-uptake system. The locus has an operonic organization and the transcriptional start site is located 292 bp upstream of hmuY. The data indicate that the regulation of the operon is iron-dependent. Interestingly, differential regulation within the operon was demonstrated, resulting in excess of the hmuYR message encoding the outer-membrane proteins when compared to the full-length transcript. In addition, the hmu Y transcript is more prevalent than the hmuR transcript. Secondary structure analysis of the hmuYRSTUV mRNA predicted the formation of several potential stem-loops in the 5′ ends of hmuR- and hmuS-specific mRNAs, consistent with the differential regulation observed. Finally, it was demonstrated that haemin binding and uptake are elevated in iron-depleted conditions and are reduced 45% and 70%, respectively, in an hmu-deficient strain when compared to the parental strain, indicating that the hmu locus plays a major role in haemin acquisition in P.gingivalis. Since homologues of the hmu locus were also found in Bacteroides fragilis, Bacteroides thetaiotaomicron and Prevotella intermedia, these findings may have implications for a better understanding of haemin acquisition in those organisms as well.
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