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Serratia marcescensis an opportunistic pathogen that commonly causes hospital-acquired infections and can utilize chitin-enriched nutrients as an alternative energy source. This study reports the identification of a chitoporin (ChiP), termedSmChiP, from the outer membrane ofS. marcescens. Sequence alignment with genetically characterized ChiPs suggests thatSmChiP is more closely related to the monomericEcChiP fromEscherichia colithan to the trimericVhChiP fromVibrio campbellii. A single crystal ofSmChiP grown under the condition 22%(w/v) PEG 8000, 0.1 Mcalcium acetate, 0.1 MMES pH 6.0 diffracted X-ray synchrotron radiation to 1.85 Å resolution.SmChiP co-crystallized with chitohexaose under the condition 19%(w/v) PEG 1500, 2 Mammonium phosphate monobasic, 0.1 MHEPES pH 7.0 diffracted X-rays to 2.70 Å resolution. Preliminary crystallographic analysis shows that bothSmChiP crystal forms contain one molecule per asymmetric unit and that they belong to the tetragonal space groupsP42212 andP41212, respectively. TheSmChiP crystal has unit-cell parametersa= 82.97,b= 82.97,c= 189.53 Å, α = β = γ = 90°, while the crystal ofSmChiP in complex with chitohexaose has unit-cell parametersa = 73.24,b= 73.24,c= 213.46 Å, α = β = γ = 90°. Initial assessment of the complex structure clearly revealed electron density for the sugar ligand. Structure determination ofSmChiP in the absence and presence of chitohexaose should reveal the molecular basis of chitin utilization byS. marcescens.

Vibrio cholerae is a Gram-negative, facultative anaerobic bacterial species that causes serious disease and can grow on various carbon sources, including chitin polysaccharides. In saltwater, its attachment to chitin surfaces not only serves as the initial step of nutrient recruitment but is also a crucial mechanism underlying cholera epidemics. In this study, we report the first characterization of a chitooligosaccharide-specific chitoporin, VcChiP, from the cell envelope of the V. cholerae type strain O1. We modeled the structure of VcChiP, revealing a trimeric cylinder that forms single channels in phospholipid bilayers. The membrane-reconstituted VcChiP channel was highly dynamic and voltage induced. Substate openings O1′, O2′, and O3′, between the fully open states O1, O2, and O3, were polarity selective, with nonohmic conductance profiles. Results of liposome-swelling assays suggested that VcChiP can transport monosaccharides, as well as chitooligosaccharides, but not other oligosaccharides. Of note, an outer-membrane porin (omp)-deficient strain of Escherichia coli expressing heterologous VcChiP could grow on M9 minimal medium supplemented with small chitooligosaccharides. These results support a crucial role of chitoporin in the adaptive survival of bacteria on chitinous nutrients. Our findings also suggest a promising means of vaccine development based on surface-exposed outer-membrane proteins and the design of novel anticholera agents based on chitooligosaccharide-mimicking analogs.

Vibrio cholerae is a Gram-negative, facultative anaerobic bacterial species that causes serious disease and can grow on various carbon sources, including chitin polysaccharides. In saltwater, its attachment to chitin surfaces not only serves as the initial step of nutrient recruitment but is also a crucial mechanism underlying cholera epidemics. In this study, we report the first characterization of a chitooligosaccharide-specific chitoporin, VcChiP, from the cell envelope of the V. cholerae type strain O1. We modeled the structure of VcChiP, revealing a trimeric cylinder that forms single channels in phospholipid bilayers. The membrane-reconstituted VcChiP channel was highly dynamic and voltage induced. Substate openings O(1)′, O(2)′, and O(3)′, between the fully open states O(1), O(2), and O(3), were polarity selective, with nonohmic conductance profiles. Results of liposome-swelling assays suggested that VcChiP can transport monosaccharides, as well as chitooligosaccharides, but not other oligosaccharides. Of note, an outer-membrane porin (omp)-deficient strain of Escherichia coli expressing heterologous VcChiP could grow on M9 minimal medium supplemented with small chitooligosaccharides. These results support a crucial role of chitoporin in the adaptive survival of bacteria on chitinous nutrients. Our findings also suggest a promising means of vaccine development based on surface-exposed outer-membrane proteins and the design of novel anticholera agents based on chitooligosaccharide-mimicking analogs.