Vinylogous Amide Analogues of Diaminopimelic Acid (DAP) as Inhibitors of Enzymes Involved in Bacterial Lysine Biosynthesis

Abstract Background: The dehydrogenase pathway and the succinylase pathway are involved in the synthesis of L-lysine in Corynebacterium glutamicum. Despite the low contribution rate to L-lysine production, the dehydrogenase pathway is favorable for its simple steps and great potentials to increase the production of L-lysine. Results: The aim of this work is to enhance the carbon flux in dehydrogenase pathway to promote L-lysine production. Firstly, the effect of ammonium (NH4+) concentration on L-lysine biosynthesis was investigated, and the results indicated that the biosynthesis of L-lysine can be promoted in high NH4+ environment. In order to reduce the requirement of NH4+, the nitrogen source regulatory protein AmtR was knocked out, resulting in an 8.5% increase in L-lysine production (i.e., 52.3±4.31 g/L). Subsequently, the dehydrogenase pathway was upregulated by blocking or weakening tetrahydrodipicolinate succinylase (DapD)-coding gene dapD and overexpressing the ddh gene to further enhance L-lysine biosynthesis. The final strain XQ-5-W4 could produce 189±8.7 g/L L-lysine with the maximum specific rate (qLys,max.) of 0.35±0.05 g/(g·h) in a 5-L jar fermenter. Conclusions: The L-lysine titer and qLys,max achieved in this study is about 25.2% and 59.1% higher than that of the original strain without enhancement of dehydrogenase pathway, respectively. The results indicated that the dehydrogenase pathway could serve as a breakthrough point to reconstruct the diaminopimelic acid (DAP) pathway and promote L-lysine production.

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Reconstruction of the Diaminopimelic Acid Pathway to Promote L-lysine Production in Corynebacterium glutamicum

International Journal of Molecular Sciences ◽

10.3390/ijms22169065 ◽

2021 ◽

Vol 22 (16) ◽

pp. 9065

Author(s):

Ning Liu ◽

Ting-Ting Zhang ◽

Zhi-Ming Rao ◽

Wei-Guo Zhang ◽

Jian-Zhong Xu

Keyword(s):

Corynebacterium Glutamicum ◽

Regulatory Protein ◽

Lysine Biosynthesis ◽

Diaminopimelic Acid ◽

Specific Rate ◽

Original Strain ◽

Lysine Production ◽

Final Strain ◽

Acid Pathway ◽

Maximum Specific Rate

The dehydrogenase pathway and the succinylase pathway are involved in the synthesis of L-lysine in Corynebacterium glutamicum. Despite the low contribution rate to L-lysine production, the dehydrogenase pathway is favorable for its simple steps and potential to increase the production of L-lysine. The effect of ammonium (NH4+) concentration on L-lysine biosynthesis was investigated, and the results indicated that the biosynthesis of L-lysine can be promoted in a high NH4+ environment. In order to reduce the requirement of NH4+, the nitrogen source regulatory protein AmtR was knocked out, resulting in an 8.5% increase in L-lysine production (i.e., 52.3 ± 4.31 g/L). Subsequently, the dehydrogenase pathway was upregulated by blocking or weakening the tetrahydrodipicolinate succinylase (DapD)-coding gene dapD and overexpressing the ddh gene to further enhance L-lysine biosynthesis. The final strain XQ-5-W4 could produce 189 ± 8.7 g/L L-lysine with the maximum specific rate (qLys,max.) of 0.35 ± 0.05 g/(g·h) in a 5-L jar fermenter. The L-lysine titer and qLys,max achieved in this study is about 25.2% and 59.1% higher than that of the original strain without enhancement of dehydrogenase pathway, respectively. The results indicated that the dehydrogenase pathway could serve as a breakthrough point to reconstruct the diaminopimelic acid (DAP) pathway and promote L-lysine production.

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Methanococci Use the Diaminopimelate Aminotransferase (DapL) Pathway for Lysine Biosynthesis

Journal of Bacteriology ◽

10.1128/jb.00172-10 ◽

2010 ◽

Vol 192 (13) ◽

pp. 3304-3310 ◽

Cited By ~ 15

Author(s):

Yuchen Liu ◽

Robert H. White ◽

William B. Whitman

Keyword(s):

Specific Activity ◽

Sequence Similarity ◽

Phylogenetic Analyses ◽

Direct Conversion ◽

Maximum Activity ◽

Lysine Biosynthesis ◽

Diaminopimelic Acid ◽

Reading Frame ◽

Cell Extracts ◽

Methanothermobacter Thermautotrophicus

ABSTRACT The pathway of lysine biosynthesis in the methanococci has not been identified previously. A variant of the diaminopimelic acid (DAP) pathway uses diaminopimelate aminotransferase (DapL) to catalyze the direct conversion of tetrahydrodipicolinate (THDPA) to ll-DAP. Recently, the enzyme DapL (MTH52) was identified in Methanothermobacter thermautotrophicus and shown to belong to the DapL1 group. Although the Methanococcus maripaludis genome lacks a gene that can be unambiguously assigned a DapL function based on sequence similarity, the open reading frame MMP1527 product shares 30% amino acid sequence identity with MTH52. A Δmmp1527 deletion mutant was constructed and found to be a lysine auxotroph, suggesting that this DapL homolog in methanococci is required for lysine biosynthesis. In cell extracts of the M. maripaludis wild-type strain, the specific activity of DapL using ll-DAP and α-ketoglutarate as substrates was 24.3 ± 2.0 nmol min−1 mg of protein−1. The gene encoding the DapL homolog in Methanocaldococcus jannaschii (MJ1391) was cloned and expressed in Escherichia coli, and the protein was purified. The maximum activity of MJ1391 was observed at 70°C and pH 8.0 to 9.0. The apparent Km s of MJ1391 for ll-DAP and α-ketoglutarate were 82.8 ± 10 μM and 0.42 ± 0.02 mM, respectively. MJ1391 was not able to use succinyl-DAP or acetyl-DAP as a substrate. Phylogenetic analyses suggested that two lateral gene transfers occurred in the DapL genes, one from the archaea to the bacteria in the DapL2 group and one from the bacteria to the archaea in the DapL1 group. These results demonstrated that the DapL pathway is present in marine methanogens belonging to the Methanococcales.

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Discovery of Heterocyclic Analogs of Diaminopimelic Acid as Promising Antibacterial Agents Through Enzyme Targeted Inhibition of Lysine Biosynthesis

Current Enzyme Inhibition ◽

10.2174/1573408014666171218150219 ◽

2018 ◽

Vol 14 (2) ◽

pp. 120-130

Author(s):

Mohd S. Shaikh ◽

Mayura A. Kale ◽

Khan Sharuk

Keyword(s):

Antibacterial Agents ◽

Lysine Biosynthesis ◽

Diaminopimelic Acid ◽

Targeted Inhibition

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Discovery of Heterocyclic Analogs of Diaminopimelic Acid as Promising Antibacterial Agents through Enzyme Targeted Inhibition of Lysine Biosynthesis

Organic Chemistry Current Research ◽

10.4172/2161-0401.1000185 ◽

2018 ◽

Vol 07 (01) ◽

Author(s):

Shaikh MS

Keyword(s):

Antibacterial Agents ◽

Lysine Biosynthesis ◽

Diaminopimelic Acid ◽

Targeted Inhibition

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Lysine biosynthesis inMethanobacterium thermoautotrophicum is by the diaminopimelic acid pathway

Current Microbiology ◽

10.1007/bf01627254 ◽

1984 ◽

Vol 10 (4) ◽

pp. 195-198 ◽

Cited By ~ 23

Author(s):

Nouna Bakhiet ◽

Fred W. Forney ◽

Donald P. Stahly ◽

Lacy Daniels

Keyword(s):

Lysine Biosynthesis ◽

Diaminopimelic Acid ◽

Acid Pathway

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Selectivity of Inhibition of N-Succinyl-l,l-Diaminopimelic Acid Desuccinylasein Bacteria: The product of dapE-gene Is Not the Target ofl-Captopril Antimicrobial Activity

Bioinorganic Chemistry and Applications ◽

10.1155/2011/306465 ◽

2011 ◽

Vol 2011 ◽

pp. 1-6 ◽

Cited By ~ 5

Author(s):

Narasimha Rao Uda ◽

Marc Creus

Keyword(s):

Antimicrobial Activity ◽

Lysine Biosynthesis ◽

Diaminopimelic Acid ◽

Antimicrobial Compounds ◽

Bacterial Strains ◽

Gram Negative ◽

Antibiotic Development ◽

Gram Positive Bacteria ◽

Good Target ◽

Peptidoglycan Cell Wall

The emergence of bacterial strains that are resistant to virtually all currently available antibiotics underscores the importance of developing new antimicrobial compounds. N-succinyl-l,l-diaminopimelic acid desuccinylase (DapE) is a metallohydrolase involved in the meso-diaminopimelate (mDAP)/lysine biosynthetic pathway necessary for lysine biosynthesis and for building the peptidoglycan cell wall. Because DapE is essential for Gram-negative and some Gram-positive bacteria, DapE has been proposed as a good target for antibiotic development. Recently,l-captopril has been suggested as a lead compound for inhibition of DapE, although its selectivity for this enzyme target in bacteria remains unclear (Gillner et al. (2009)). Here, we tested the selectivity ofl-captopril against DapEin bacteria. Since DapE knockout strains of gram-negative bacteria are viable upon chemical supplementation with mDAP, we reasoned that the antimicrobial activity of compounds targeting DapE should be abolished in mDAP-containing media. Althoughl-captopril had modest antimicrobial activity inEscherichia coliand inSalmonella enterica, to our surprise, inhibition of bacterial growth was independent both of mDAP supplementation and DapE over-expression. We conclude that DapE is not the main target ofl-captopril inhibition in these bacteria. The methods implemented here will be useful for screening DapE-selective antimicrobial compounds directly in bacterial cultures.

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