Identification and characterization of a Bacteroides gene, csuF, which encodes an outer membrane protein that is essential for growth on chondroitin sulfate.
Bacteroides thetaiotaomicron can utilize a variety of polysaccharides, including charged mucopolysaccharides such as chondroitin sulfate (CS) and hyaluronic acid (HA). Since the enzymes (chondroitin lyases I and II) that catalyze the first step in breakdown of CS and HA are located in the periplasm, we had proposed that the first step in utilization of these polysaccharides was binding to one or more outer membrane proteins followed by translocation into the periplasm, but no such outer membrane proteins had been shown to play a role in CS or HA utilization. Previously we have isolated a transposon-generated mutant, CS4, which was unable to grow on CS or HA but retained the ability to grow on disaccharide components of CS. This phenotype suggested that the mutation in CS4 either blocked the transport of the mucopolysaccharides into the periplasmic space or blocked the depolymerization of the mucopolysaccharides into disaccharides. We have mapped the CS4 mutation to a single gene, csuF, which is capable of encoding a protein of 1,065 amino acids and contains a consensus signal sequence. Although CsuF had a predicted molecular weight and pI similar to those of chondroitin lyases, it did not show significant sequence similarity to the Bacteroides chondroitin lyase II, a Proteus chondroitin ABC lyase, or two hyaluronidases from Clostridium perfringens and Streptococcus pyogenes, nor was any CS-degrading enzyme activity associated with csuF expression in Bacteroides species or Escherichia coli. The deduced amino acid sequence of CsuF exhibited features suggestive of an outer membrane protein. We obtained antibodies to CsuF and demonstrated that the protein is located in the outer membrane. This is the first evidence that a nonenzymatic outer membrane protein is essential for utilization of CS and HA.
Cheng
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Q
Department of Microbiology, University of Illinois, Urbana 61801, USA.
Yu
M C
MC
Reeves
A R
AR
Salyers
A A
AA
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D001439:Bacteroides / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D002451:Cell Compartmentation; D002462:Cell Membrane / Q000737:chemistry; D002808:Chondroitin Lyases / Q000235:genetics; D002809:Chondroitin Sulfates / Q000378:metabolism; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial / Q000235:genetics*; D006025:Glycosaminoglycans / Q000378:metabolism*; D006820:Hyaluronic Acid / Q000378:metabolism; D006821:Hyaluronoglucosaminidase / Q000235:genetics; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D016366:Open Reading Frames; D021382:Protein Sorting Signals / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis
D001425:Bacterial Outer Membrane Proteins; C094420:CsuF protein, bacteria; D006025:Glycosaminoglycans; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D006820:Hyaluronic Acid; D002809:Chondroitin Sulfates; D006821:Hyaluronoglucosaminidase; D002808:Chondroitin Lyases