Porin OmpP2 of Haemophilus influenzae shows specificity for nicotinamide-derived nucleotide substrates.
Haemophilus influenzae has an absolute requirement for NAD (factor V) because it lacks all biosynthetic enzymes necessary for de novo synthesis of that cofactor. Therefore, growth in vitro requires the presence of NAD itself, NMN, or nicotinamide riboside (NR). To address uptake abilities of these compounds, we investigated outer membrane proteins. By analyzing ompP2 knockout mutants, we found that NAD and NMN uptake was prevented, whereas NR uptake was not. Through investigation of the properties of purified OmpP2 in artificial lipid membrane systems, the substrate specificity of OmpP2 for NAD and NMN was determined, with KS values of approximately 8 and 4mm, respectively, in 0.1 m KCl, whereas no interaction was detected for the nucleoside NR and other purine or pyrimidine nucleotide or nucleoside species. Based on our analysis, we assume that an intrinsic binding site within OmpP2 exists that facilitates diffusion of these compounds across the outer membrane, recognizing carbonyl and exposed phosphate groups. Because OmpP2 was formerly described as a general diffusion porin, an additional property of acting as a facilitator for nicotinamide-based nucleotide transport may have evolved to support and optimize utilization of the essential cofactor sources NAD and NMN in H. influenzae.
Andersen
Christian
C
Lehrstuhl für Biotechnologie, Biozentrum der Universität Würzburg, Am Hubland, Germany.
Maier
Elke
E
Kemmer
Gabrielle
G
Blass
Julia
J
Hilpert
Anna-Karina
AK
Benz
Roland
R
Reidl
Joachim
J
D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics; D001692:Biological Transport; D006193:Haemophilus influenzae / Q000201:enzymology; D009154:Mutation; D009243:NAD / Q000378:metabolism*; D009537:Nicotinamide Mononucleotide / Q000378:metabolism*; D011485:Protein Binding; D013379:Substrate Specificity
D001425:Bacterial Outer Membrane Proteins; C074703:outer membrane protein P2, Haemophilus influenzae type b; D009243:NAD; D009537:Nicotinamide Mononucleotide