Vaccine target and carrier molecule nontypeable Haemophilus influenzae protein D dimerizes like the close Escherichia coli GlpQ homolog but unlike other known homolog dimers

We have determined the 1.8 Å X‐ray crystal structure of nonlipidated (i.e., N‐terminally truncated) nontypeable Haemophilus influenzae (NTHi; H. influenzae) protein D. Protein D exists on outer membranes of H. influenzae strains and acts as a virulence factor that helps invade human cells. Protein D...

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Published in:	Proteins, structure, function, and bioinformatics Vol. 91; no. 2; pp. 161 - 170
Main Authors:	Jones, Seth P., Cook, Kali H., Holmquist, Melody L., Almekinder, Liam J., Delaney, Annie M., Charles, Ryhl, Labbe, Natalie, Perdue, Janai, Jackson, Niaya, Pichichero, Michael E., Kaur, Ravinder, Michel, Lea V., Gleghorn, Michael L.
Format:	Journal Article
Language:	English
Published:	Hoboken, USA John Wiley & Sons, Inc 01-02-2023 Wiley Subscription Services, Inc
Subjects:	Amino acid sequence Animal models Antigens Carrier Proteins - genetics Chronic obstructive pulmonary disease COPD Crystal lattices Crystal structure Dimerization E coli Escherichia coli Escherichia coli - genetics Glycerol Glycerophosphodiester phosphodiesterase Haemophilus influenzae Haemophilus influenzae - genetics Homology Hydrolases Interfaces Intermediates Kinases Lung diseases Mass spectrometry Mass spectroscopy Monomers nontypeable haemophilus influenzae Obstructive lung disease Otitis media Outer membranes Phosphodiesterase Polysaccharides Protein D Protein structure Proteins Saccharides Size exclusion chromatography Structural analysis Substrates Vaccines Virulence factors nontypeable haemophilus influenzae otitis media glycerol COPD glycerophosphodiester phosphodiesterase
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Summary:	We have determined the 1.8 Å X‐ray crystal structure of nonlipidated (i.e., N‐terminally truncated) nontypeable Haemophilus influenzae (NTHi; H. influenzae) protein D. Protein D exists on outer membranes of H. influenzae strains and acts as a virulence factor that helps invade human cells. Protein D is a proven successful antigen in animal models to treat obstructive pulmonary disease (COPD) and otitis media (OM), and when conjugated to polysaccharides also has been used as a carrier molecule for human vaccines, for example in GlaxoSmithKline Synflorix™. NTHi protein D shares high sequence and structural identify to the Escherichia coli (E. coli) glpQ gene product (GlpQ). E. coli GlpQ is a glycerophosphodiester phosphodiesterase (GDPD) with a known dimeric structure in the Protein Structural Database, albeit without an associated publication. We show here that both structures exhibit similar homodimer organization despite slightly different crystal lattices. Additionally, we have observed both the presence of weak dimerization and the lack of dimerization in solution during size exclusion chromatography (SEC) experiments yet have distinctly observed dimerization in native mass spectrometry analyses. Comparison of NTHi protein D and E. coli GlpQ with other homologous homodimers and monomers shows that the E. coli and NTHi homodimer interfaces are distinct. Despite this distinction, NTHi protein D and E. coli GlpQ possess a triose‐phosphate isomerase (TIM) barrel domain seen in many of the other homologs. The active site of NTHi protein D is located near the center of this TIM barrel. A putative glycerol moiety was modeled in two different conformations (occupancies) in the active site of our NTHi protein D structure and we compared this to ligands modeled in homologous structures. Our structural analysis should aid in future efforts to determine structures of protein D bound to substrates, analog intermediates, and products, to fully appreciate this reaction scheme and aiding in future inhibitor design.
Bibliography:	Funding information R21 AI153936‐01 NIDCD (PI: Kaur) and Rochester Regional Health (Pichichero) Seth P. Jones and Kali H. Cook Co‐first authors. ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	0887-3585 1097-0134
DOI:	10.1002/prot.26418