Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is well known for its involvement in numerous non-metabolic processes inside mammalian cells. Alternative functions of prokaryotic GAPDH are mainly deduced from its extracellular localization ability to bind to selected host proteins. Data on its participation in intracellular bacterial processes are scarce as there has been to date only one study dealing with this issue. We previously have reported several points of evidence that the GAPDH homolog of Francisella tularensis GapA might also exert additional non-enzymatic functions. Following on from our earlier observations we decided to identify GapA's interacting partners within the bacterial proteome to explore its new roles at intracellular level. The quantitative proteomics approach based on stable isotope labeling of amino acids in cell culture (SILAC) in combination with affinity purification mass spectrometry enabled us to identify 18 proteins potentially interacting with GapA. Six of those interactions were further confirmed by alternative methods. Half of the identified proteins were involved in non-metabolic processes. Further analysis together with quantitative label-free comparative analysis of proteomes isolated from the wild-type strain strain with deleted gapA gene suggests that GapA is implicated in DNA repair processes. Absence of GapA promotes secretion of its most potent interaction partner the hypothetical protein with peptidase propeptide domain (PepSY) thereby indicating that it impacts on subcellular distribution of some proteins.

• Keywords
• Francisella tularensis, SILAC, glyceraldehyde-3-phosphate dehydrogenase, multifunctional enzyme, protein–protein interaction,
• Publication type
• Journal Article MeSH

Bacterial proteins exhibiting two or more unrelated functions, referred to as moonlighting proteins, are suggested to contribute to full virulence manifestation in pathogens. An expanding number of published studies have revealed the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) to be a multitasking protein with virulence impact in a number of pathogenic bacteria. This protein can be detected on the bacterial surface or outside the bacterial cell, where it interacts with host proteins. In this way, GAPDH is able to modulate various pathogenic processes. Moreover, it has been shown to be involved in non-enzymatic processes inside the bacterial cell. In this mini review, we summarize main findings concerning the multiple localization and protein interactions of GAPDH derived from bacterial pathogens of humans. We also briefly discuss problems associated with using GAPDH as a vaccine antigen and endeavor to inspire further research to fill gaps in the existing knowledge.

• Keywords
• glyceraldehyde-3-phosphate dehydrogenase, localization, moonlighting proteins, pathogenic bacteria, protein-protein interaction (PPI),
• MeSH
• Bacteria enzymology   pathogenicity   MeSH
• Bacterial Infections microbiology   prevention & control   MeSH
• Bacterial Proteins metabolism   MeSH
• Bacterial Vaccines immunology   MeSH
• Glyceraldehyde-3-Phosphate Dehydrogenases immunology   metabolism   MeSH
• Humans MeSH
• Proteins metabolism   MeSH
• Protein Binding MeSH
• Virulence MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Names of Substances
• Bacterial Proteins MeSH
• Bacterial Vaccines MeSH
• Glyceraldehyde-3-Phosphate Dehydrogenases MeSH
• Proteins MeSH

It is becoming increasingly evident that a high degree of regulation is involved in the protein synthesis machinery entailing more interacting regulatory factors. A multitude of proteins have been identified recently which show regulatory function upon binding to the ribosome. Here, we identify tight association of a metabolic protein aldehyde-alcohol dehydrogenase E (AdhE) with the E. coli 70S ribosome isolated from cell extract under low salt wash conditions. Cryo-EM reconstruction of the ribosome sample allows us to localize its position on the head of the small subunit, near the mRNA entrance. Our study demonstrates substantial RNA unwinding activity of AdhE which can account for the ability of ribosome to translate through downstream of at least certain mRNA helices. Thus far, in E. coli, no ribosome-associated factor has been identified that shows downstream mRNA helicase activity. Additionally, the cryo-EM map reveals interaction of another extracellular protein, outer membrane protein C (OmpC), with the ribosome at the peripheral solvent side of the 50S subunit. Our result also provides important insight into plausible functional role of OmpC upon ribosome binding. Visualization of the ribosome purified directly from the cell lysate unveils for the first time interactions of additional regulatory proteins with the ribosome.

• MeSH
• Aldehyde Oxidoreductases chemistry   metabolism   MeSH
• Alcohol Dehydrogenase chemistry   metabolism   MeSH
• Escherichia coli metabolism   MeSH
• Protein Conformation MeSH
• Models, Molecular MeSH
• Porins chemistry   metabolism   MeSH
• Escherichia coli Proteins chemistry   metabolism   MeSH
• Ribosomes chemistry   genetics   metabolism   MeSH
• Protein Binding MeSH
• Binding Sites MeSH
• Structure-Activity Relationship MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• adhE protein, E coli MeSH Browser
• Aldehyde Oxidoreductases MeSH
• Alcohol Dehydrogenase MeSH
• OmpC protein MeSH Browser
• Porins MeSH
• Escherichia coli Proteins MeSH