scholarly journals Comparative Genomics of Ethanolamine Utilization

2009 ◽  
Vol 191 (23) ◽  
pp. 7157-7164 ◽  
Author(s):  
Olga Tsoy ◽  
Dmitry Ravcheev ◽  
Arcady Mushegian
Keyword(s):  
Computational Analysis ◽  
Catalytic Mechanism ◽  
Food Poisoning ◽  
Structural Components ◽  
Carbon And Nitrogen ◽  
Dna Motif ◽  
Serovar Typhimurium ◽  
Tim Barrel ◽  
Per Se ◽  
Horizontal Transfers

ABSTRACT Ethanolamine can be used as a source of carbon and nitrogen by phylogenetically diverse bacteria. Ethanolamine-ammonia lyase, the enzyme that breaks ethanolamine into acetaldehyde and ammonia, is encoded by the gene tandem eutBC. Despite extensive studies of ethanolamine utilization in Salmonella enterica serovar Typhimurium, much remains to be learned about EutBC structure and catalytic mechanism, about the evolutionary origin of ethanolamine utilization, and about regulatory links between the metabolism of ethanolamine itself and the ethanolamine-ammonia lyase cofactor adenosylcobalamin. We used computational analysis of sequences, structures, genome contexts, and phylogenies of ethanolamine-ammonia lyases to address these questions and to evaluate recent data-mining studies that have suggested an association between bacterial food poisoning and the diol utilization pathways. We found that EutBC evolution included recruitment of a TIM barrel and a Rossmann fold domain and their fusion to N-terminal α-helical domains to give EutB and EutC, respectively. This fusion was followed by recruitment and occasional loss of auxiliary ethanolamine utilization genes in Firmicutes and by several horizontal transfers, most notably from the firmicute stem to the Enterobacteriaceae and from Alphaproteobacteria to Actinobacteria. We identified a conserved DNA motif that likely represents the EutR-binding site and is shared by the ethanolamine and cobalamin operons in several enterobacterial species, suggesting a mechanism for coupling the biosyntheses of apoenzyme and cofactor in these species. Finally, we found that the food poisoning phenotype is associated with the structural components of metabolosome more strongly than with ethanolamine utilization genes or with paralogous propanediol utilization genes per se.

scholarly journals A Mannose Family Phosphotransferase System Permease and Associated Enzymes Are Required for Utilization of Fructoselysine and Glucoselysine in Salmonella enterica Serovar Typhimurium

2015 ◽  
Vol 197 (17) ◽  
pp. 2831-2839 ◽  
Author(s):  
Katherine A. Miller ◽  
Robert S. Phillips ◽  
Paul B. Kilgore ◽  
Grady L. Smith ◽  
Timothy R. Hoover
Keyword(s):  
Nitrogen Source ◽  
Nitrogen Sources ◽  
Maillard Reaction Products ◽  
Reaction Products ◽  
Phosphotransferase System ◽  
Carbon And Nitrogen Sources ◽  
Content Type ◽  
Carbon And Nitrogen ◽  
Serovar Typhimurium ◽  
Food Borne Illness

ABSTRACTSalmonella entericserovar Typhimurium, a major cause of food-borne illness, is capable of using a variety of carbon and nitrogen sources. Fructoselysine and glucoselysine are Maillard reaction products formed by the reaction of glucose or fructose, respectively, with the ε-amine group of lysine. We report here thatS. Typhimurium utilizes fructoselysine and glucoselysine as carbon and nitrogen sources via a mannose family phosphotransferase (PTS) encoded bygfrABCD(glucoselysine/fructoselysine PTS components EIIA, EIIB, EIIC, and EIID; locus numbers STM14_5449 to STM14_5454 inS. Typhimurium 14028s). Genes coding for two predicted deglycases within thegfroperon,gfrEandgfrF, were required for growth with glucoselysine and fructoselysine, respectively. GfrF demonstrated fructoselysine-6-phosphate deglycase activity in a coupled enzyme assay. The biochemical and genetic analyses were consistent with a pathway in which fructoselysine and glucoselysine are phosphorylated at the C-6 position of the sugar by the GfrABCD PTS as they are transported across the membrane. The resulting fructoselysine-6-phosphate and glucoselysine-6-phosphate subsequently are cleaved by GfrF and GfrE to form lysine and glucose-6-phosphate or fructose-6-phosphate. Interestingly, althoughS. Typhimurium can use lysine derived from fructoselysine or glucoselysine as a sole nitrogen source, it cannot use exogenous lysine as a nitrogen source to support growth. Expression ofgfrABCDEFwas dependent on the alternative sigma factor RpoN (σ54) and an RpoN-dependent LevR-like activator, which we designated GfrR.IMPORTANCESalmonellaphysiology has been studied intensively, but there is much we do not know regarding the repertoire of nutrients these bacteria are able to use for growth. This study shows that a previously uncharacterized PTS and associated enzymes function together to transport and catabolize fructoselysine and glucoselysine. Knowledge of the range of nutrients thatSalmonellautilizes is important, as it could lead to the development of new strategies for reducing the load ofSalmonellain food animals, thereby mitigating its entry into the human food supply.

scholarly journals Loss of Ethanolamine Utilization in Enterococcus faecalis Increases Gastrointestinal Tract Colonization

mBio ◽  
2018 ◽  
Vol 9 (3) ◽  
Author(s):  
Karan Gautam Kaval ◽  
Kavindra V. Singh ◽  
Melissa R. Cruz ◽  
Sruti DebRoy ◽  
Wade C. Winkler ◽  
...  
Keyword(s):  
Enterococcus Faecalis ◽  
Opposite Effect ◽  
Wild Type Strain ◽  
Food Poisoning ◽  
Wild Type ◽  
Gi Tract ◽  
Human Gut ◽  
Content Type ◽  
Serovar Typhimurium ◽  
High Concentrations

ABSTRACT Enterococcus faecalis is paradoxically a dangerous nosocomial pathogen and a normal constituent of the human gut microbiome, an environment rich in ethanolamine. E. faecalis carries the eut (ethanolamine utilization) genes, which enable the catabolism of ethanolamine (EA) as a valuable source of carbon and/or nitrogen. EA catabolism was previously shown to contribute to the colonization and growth of enteric pathogens, such as Salmonella enterica serovar Typhimurium and enterohemorrhagic Escherichia coli (EHEC), in the gut environment. We tested the ability of eut mutants of E. faecalis to colonize the gut using a murine model of gastrointestinal (GI) tract competition and report the surprising observation that these mutants outcompete the wild-type strain. IMPORTANCE Some bacteria that are normal, harmless colonizers of the human body can cause disease in immunocompromised patients, particularly those that have been heavily treated with antibiotics. Therefore, it is important to understand the factors that promote or negate these organisms’ ability to colonize. Previously, ethanolamine, found in high concentrations in the GI tract, was shown to promote the colonization and growth of bacteria associated with food poisoning. Here, we report the surprising, opposite effect of ethanolamine utilization on the commensal colonizer E. faecalis , namely, that loss of this metabolic capacity made it a better colonizer.

scholarly journals Structural Insights into the Molecular Evolution of the Archaeal Exo-β-d-Glucosaminidase

2019 ◽  
Vol 20 (10) ◽  
pp. 2460
Author(s):  
Shouhei Mine ◽  
Masahiro Watanabe
Keyword(s):  
Molecular Evolution ◽  
Primary Structure ◽  
Active Site ◽  
Catalytic Mechanism ◽  
Thermostable Enzyme ◽  
Crystallographic Analysis ◽  
Evolutionary Pathway ◽  
Chitosan Oligosaccharides ◽  
Tim Barrel ◽  
Structural Insights

The archaeal exo-β-d-glucosaminidase (GlmA), a thermostable enzyme belonging to the glycosidase hydrolase (GH) 35 family, hydrolyzes chitosan oligosaccharides into monomer glucosamines. GlmA is a novel enzyme in terms of its primary structure, as it is homologous to both GH35 and GH42 β-galactosidases. The catalytic mechanism of GlmA is not known. Here, we summarize the recent reports on the crystallographic analysis of GlmA. GlmA is a homodimer, with each subunit comprising three distinct domains: a catalytic TIM-barrel domain, an α/β domain, and a β1 domain. Surprisingly, the structure of GlmA presents features common to GH35 and GH42 β-galactosidases, with the domain organization resembling that of GH42 β-galactosidases and the active-site architecture resembling that of GH35 β-galactosidases. Additionally, the GlmA structure also provides critical information about its catalytic mechanism, in particular, on how the enzyme can recognize glucosamine. Finally, we postulate an evolutionary pathway based on the structure of an ancestor GlmA to extant GH35 and GH42 β-galactosidases.

scholarly journals Crystal structures of γ-glutamylmethylamide synthetase provide insight into bacterial metabolism of oceanic monomethylamine

2020 ◽  
pp. jbc.RA120.015952
Author(s):  
Ning Wang ◽  
Xiu-Lan Chen ◽  
Chao Gao ◽  
Ming Peng ◽  
Peng Wang ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Catalytic Mechanism ◽  
Structural Information ◽  
Trace Gas ◽  
Microbial Genomes ◽  
Carbon And Nitrogen ◽  
Tetrahedral Intermediate ◽  
Surface Ocean ◽  
Biochemical Analyses ◽  
Global Carbon

Monomethylamine (MMA) is an important climate-active oceanic trace gas and ubiquitous in the oceans. The γ-glutamylmethylamide synthetase (GmaS) catalyzes the conversion of MMA to γ-glutamylmethylamide (GMA), the first step in MMA metabolism in many marine bacteria. The gmaS gene occurs in ~23% of microbial genomes in the surface ocean and is a validated biomarker to detect MMA-utilizing bacteria. However, the catalytic mechanism of GmaS has not been studied due to the lack of structural information. Here, the GmaS from Rhodovulum sp. 12E13 (RhGmaS) was characterized, and the crystal structures of apo-RhGmaS and RhGmaS with different ligands in five states were solved. Based on structural and biochemical analyses, the catalytic mechanism of RhGmaS was explained. ATP is first bound in RhGmaS, leading to a conformational change of a flexible loop (Lys287-Ile305), which is essential for the subsequent binding of glutamate. During the catalysis of RhGmaS, the residue Arg312 participates in polarizing the γ-phosphate of ATP and in stabilizing the γ-glutamyl phosphate intermediate; Asp177 is responsible for the deprotonation of MMA, assisting the attack of MMA on γ-glutamyl phosphate to produce a tetrahedral intermediate; and Glu186 acts as a catalytic base to abstract a proton from the tetrahedral intermediate to finally generate GMA. Sequence analysis suggested that the catalytic mechanism of RhGmaS proposed in this study has universal significance in bacteria containing GmaS. Our results provide novel insights into MMA metabolism, contributing to a better understanding of MMA catabolism in global carbon and nitrogen cycles.

scholarly journals A Novel Deaminase Involved in Chloronitrobenzene and Nitrobenzene Degradation with Comamonas sp. Strain CNB-1

2007 ◽  
Vol 189 (7) ◽  
pp. 2677-2682 ◽  
Author(s):  
Lei Liu ◽  
Jian-Feng Wu ◽  
Ying-Fei Ma ◽  
Sheng-Yue Wang ◽  
Guo-Ping Zhao ◽  
...  
Keyword(s):  
Amino Acids ◽  
Bacterial Growth ◽  
Bradyrhizobium Japonicum ◽  
Catalytic Mechanism ◽  
Rhodopseudomonas Palustris ◽  
Hydroxyl Group ◽  
Hypothetical Proteins ◽  
Terminal Sequence ◽  
Significant Similarity ◽  
Carbon And Nitrogen

ABSTRACT Comamonas sp. strain CNB-1 degrades nitrobenzene and chloronitrobenzene via the intermediates 2-aminomuconate and 2-amino-5-chloromuconate, respectively. Deamination of these two compounds results in the release of ammonia, which is used as a source of nitrogen for bacterial growth. In this study, a novel deaminase was purified from Comamonas strain CNB-1, and the gene (cnbZ) encoding this enzyme was cloned. The N-terminal sequence and peptide fingerprints of this deaminase were determined, and BLAST searches revealed no match with significant similarity to any functionally characterized proteins. The purified deaminase is a monomer (30 kDa), and its V max values for 2-aminomuconate and 2-amino-5-chloromuconate were 147 μmol·min−1·mg−1 and 196 μmol·min−1·mg−1, respectively. Its catalytic products from 2-aminomuconate and 2-amino-5-chloromuconate were 2-hydroxymuconate and 2-hydroxy-5-chloromuconate, respectively, which are different from those previously reported for the deaminases of Pseudomonas species. In the catalytic mechanism proposed, the α-carbon and nitrogen atoms (of both 2-aminomuconate and 2-amino-5-chloromuconate) were simultaneously attacked by a hydroxyl group and a proton, respectively. Homologs of cnbZ were identified in the genomes of Bradyrhizobium japonicum, Rhodopseudomonas palustris, and Roseiflexus sp. strain RS-1; these genes were previously annotated as encoding hypothetical proteins of unknown function. It is concluded that CnbZ represents a novel enzyme that deaminates xenobiotic compounds and/or α-amino acids.

scholarly journals Characterisation of IncI1 plasmids associated with change of phage type in isolates of Salmonella enterica serovar Typhimurium

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Lester Hiley ◽  
Rikki M. A. Graham ◽  
Amy V. Jennison
Keyword(s):  
Significant Variation ◽  
Genetic Recombination ◽  
Phage Type ◽  
Food Poisoning ◽  
Phage Types ◽  
Gene Sets ◽  
Acid Protein ◽  
Serovar Typhimurium ◽  
New Perspective ◽  
Bacteriophage Bf23

Abstract Background Acquisition of IncI1 plasmids by members of the Enterobacteriaceae sometimes leads to transfer of antimicrobial resistance and colicinogeny as well as change of phage type in Salmonella Typhimurium. Isolates of S. Typhimurium from a 2015 outbreak of food poisoning were found to contain an IncI1 plasmid implicated in change of phage type from PT135a to U307 not previously reported. The origin of the changes of phage type associated with this IncI1 plasmid was investigated. In addition, a comparison of its gene composition with that of IncI1 plasmids found in local isolates of S. Typhimurium typed as U307 from other times was undertaken. This comparison was extended to IncI1 plasmids in isolates of phage types PT6 and PT6 var. 1 which are thought to be associated with acquisition of IncI1 plasmids. Results Analysis of IncI1 plasmids from whole genome sequencing of isolates implicated a gene coding for a 1273 amino acid protein present only in U307 isolates as the likely source of change of phage type. The IncI1 plasmids from PT6 and PT6 var. 1 isolates all had the ibfA gene present in IncI1 plasmid R64. This gene inhibits growth of bacteriophage BF23 and was therefore the possible source of change of phage type. A fuller comparison of the genetic composition of IncI1 plasmids from U307 isolates and PT6 and PT6 var. 1 isolates along with two IncI1 plasmids from S. Typhimurium isolates not showing change of phage type was undertaken. Plasmids were classified as either ‘Delta’ or ‘Col’ IncI1 plasmids according to whether genes between repZ and the rfsF site showed high identity to genes in the same location in R64 or ColIb-P9 plasmids respectively. Comparison of the tra gene sets and the pil gene sets across the range of sequenced plasmids identified Delta and Col plasmids with almost identical sequences for both sets of genes. This indicated a genetic recombination event leading to a switch between Delta and Col gene sets at the rfsF site. Comparisons of other gene sets showing significant variation among the sequenced plasmids are reported. Searches of the NCBI GenBank database using DNA and protein sequences of interest from the sequenced plasmids identified global IncI1 plasmids with extensive regions showing 99 to 100% identity to some of the plasmids sequenced in this study indicating evidence for widespread distribution of these plasmids. Conclusion Two genes possibly associated with change of phage type were identified in IncI1 plasmids. IncI1 plasmids were classified as either ‘Delta’ or ‘Col’ plasmids and other sequences of significant variation among these plasmids were identified. This study offers a new perspective on the understanding of the gene composition of IncI1 plasmids. The sequences of newly sequenced IncI1 plasmids could be compared against the regions of significant sequence variation identified in this study to understand better their overall gene composition and relatedness to other IncI1 plasmids in the databases.

scholarly journals Current state of and need for enzyme engineering of 2-deoxy-D-ribose 5-phosphate aldolases and its impact

2021 ◽  
Author(s):  
Juha Rouvinen ◽  
Martina Andberg ◽  
Johan Pääkkönen ◽  
Nina Hakulinen ◽  
Anu Koivula
Keyword(s):  
Protein Engineering ◽  
Catalytic Mechanism ◽  
Cascade Reactions ◽  
Single Step ◽  
Enzyme Engineering ◽  
Enzymatic Reactions ◽  
Tim Barrel ◽  
The Stability

Abstract Deoxyribose-5-phosphate aldolases (DERAs, EC 4.1.2.4) are acetaldehyde-dependent, Class I aldolases catalyzing in nature a reversible aldol reaction between an acetaldehyde donor (C2 compound) and glyceraldehyde-3-phosphate acceptor (C3 compound, C3P) to generate deoxyribose-5-phosphate (C5 compound, DR5P). DERA enzymes have been found to accept also other types of aldehydes as their donor, and in particular as acceptor molecules. Consequently, DERA enzymes can be applied in C–C bond formation reactions to produce novel compounds, thus offering a versatile biocatalytic alternative for synthesis. DERA enzymes, found in all kingdoms of life, share a common TIM barrel fold despite the low overall sequence identity. The catalytic mechanism is well-studied and involves formation of a covalent enzyme-substrate intermediate. A number of protein engineering studies to optimize substrate specificity, enzyme efficiency, and stability of DERA aldolases have been published. These have employed various engineering strategies including structure-based design, directed evolution, and recently also machine learning–guided protein engineering. For application purposes, enzyme immobilization and usage of whole cell catalysis are preferred methods as they improve the overall performance of the biocatalytic processes, including often also the stability of the enzyme. Besides single-step enzymatic reactions, DERA aldolases have also been applied in multi-enzyme cascade reactions both in vitro and in vivo. The DERA-based applications range from synthesis of commodity chemicals and flavours to more complicated and high-value pharmaceutical compounds. Key points • DERA aldolases are versatile biocatalysts able to make new C–C bonds. • Synthetic utility of DERAs has been improved by protein engineering approaches. • Computational methods are expected to speed up the future DERA engineering efforts. Graphical abstract

scholarly journals Detection and quantification of Salmonella in fresh vegetables in Perak, Malaysia

Food Research ◽  
2019 ◽  
Vol 4 (2) ◽  
pp. 441-448 ◽  
Author(s):  
Seow Hoon Saw ◽  
J.L. Mak ◽  
M.H. Tan ◽  
S.T. Teo ◽  
T.Y. Tan ◽  
...  
Keyword(s):  
Pathogenic Bacteria ◽  
Food Poisoning ◽  
Leafy Vegetables ◽  
Most Probable Number ◽  
Salmonella Spp ◽  
Probable Number ◽  
Increased Risk ◽  
Fresh Vegetables ◽  
Serovar Typhimurium ◽  
Wet Market

The eating of fresh and minimally processed vegetables is getting popular among Malaysians. This trend poses an increased risk of food poisoning associated with the consumption of fresh produce contaminated with pathogenic bacteria. Salmonellosis is a foodborne disease caused by several non-typhoidal Salmonella enterica serovars, predominantly serovars Enteritidis and Typhimurium. The present study aimed to determine the prevalence of Salmonella spp., S. enterica serovar Enteritidis and S. enterica serovar Typhimurium in fresh leafy vegetables such as cabbages (n = 40), lettuces (n = 20), and fruit vegetables such as tomatoes (n = 40), carrots (n = 40) and cucumbers (n = 40), which were sold by three different hypermarkets and a wet market in Kampar, Perak, Malaysia. The study was performed over a period of 13 months (January 2018 to January 2019). A combination of most probable number-multiplex polymerase chain reaction (MPN-mPCR) method was used to quantify the concentrations of Salmonella spp., S. enterica serovar Enteritidis and S. enterica serovar Typhimurium in the examined samples. The results of this study demonstrated that of the vegetables tested, tomatoes, carrots and lettuces were not contaminated by Salmonella spp., S. enterica serovar Enteritidis and S. enterica serovar Typhimurium. However, the presence of Salmonella spp. was detected in 3.3% of cabbages from the hypermarket, with estimated microbial loads ranging from <3.0 MPN/g to 15.0 MPN/g. On the other hand, S. enterica serovar Typhimurium was detected in 10.0% of the cucumbers from hypermarkets and 20% of them from the wet market. Their microbial loads were ranging from <3.0 MPN/g to >1,100 MPN/g. This indicated that cabbages and cucumbers could be the potential sources of salmonellosis. Therefore, the monitoring of food safety and hygienic practices should be strictly enforced by relevant government agencies to avoid potential poisoning by foodborne pathogens.

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