Differential response of orthologous l,l-diaminopimelate aminotransferases (DapL) to enzyme inhibitory antibiotic lead compounds

Abstract

L,L-Diaminopimelate aminotransferase (DapL) is an enzyme required for the biosynthesis of meso-diaminopimelate (m-DAP) and L-lysine (Lys) in some bacteria and photosynthetic organisms. m-DAP and Lys are both involved in the synthesis of peptidoglycan (PG) and protein synthesis. DapL is found in specific eubacterial and archaeal lineages, in particular in several groups of pathogenic bacteria such as Leptospira interrogans (LiDapL), the soil/water bacterium Verrucomicrobium spinosum (VsDapL) and the alga Chlamydomonas reinhardtii (CrDapL). Here we present the first comprehensive inhibition study comparing the kinetic activity of DapL orthologs using previously active small molecule inhibitors formerly identified in a screen with the DapL of Arabidopsis thaliana (AtDapL), a flowering plant. Each inhibitor is derived from one of four classes with different central structural moieties: a hydrazide, a rhodanine, a barbiturate, or a thiobarbituate functionality. The results show that all five compounds tested were effective at inhibiting the DapL orthologs. LiDapL and AtDapL showed similar patterns of inhibition across the inhibitor series, whereas the VsDapL and CrDapL inhibition patterns were different from that of LiDapL and AtDapL. CrDapL was found to be insensitive to the hydrazide (IC₅₀ >200 μM). VsDapL was found to be the most sensitive to the barbiturate and thiobarbiturate containing inhibitors (IC₅₀ ∼5 μM). Taken together, the data shows that the homologs have differing sensitivities to the inhibitors with IC₅₀ values ranging from 4.7 to 250 μM. In an attempt to understand the basis for these differences the four enzymes were modeled based on the known structure of AtDapL. Overall, it was found that the enzyme active sites were conserved, although the second shell of residues close to the active site were not. We conclude from this that the altered binding patterns seen in the inhibition studies may be a consequence of the inhibitors forming additional interactions with residues proximal to the active site, or that the inhibitors may not act by binding to the active site. Compounds that are specific for DapL could be potential biocides (antibiotic, herbicide or algaecide) that are nontoxic to animals since animals do not contain the enzymes necessary for PG or Lys synthesis. This study provides important information to expand our current understanding of the structure/activity relationship of DapL and putative inhibitors that are potentially useful for the design and or discovery of novel biocides.

Mechanisms on Boron-Induced Alleviation of Aluminum-Toxicity in Citrus grandis Seedlings at a Transcriptional Level Revealed by cDNA-AFLP Analysis

“…LL -diaminopimelate aminotransferase is an enzyme involved in meso -diaminopimelate, a precursor of cell wall peptidoglycan and l -lysine in plants [ 51 ]. Tyrosine transaminase (also known as tyrosine aminotransferase) catalyzes the conversion of tyrosine to 4-hydroxyphenylpyruvic acid, a precursor for homogenetisic acid, plastoquinones and tocopherols, the latter of which function as radical scavengers and protect the plants against various stresses [ 52 ].…”

Analysis of drug binding pockets and repurposing opportunities for twelve essential enzymes of ESKAPE pathogens

“…For illustration, the effect of active inhibitors on DapL orthologs like Arabidopsis thaliana (AtDapL), a flowering plant, Leptospira interrogans (LiDapL), the soil/water bacterium Verrucomicrobium spinosum (VsDapL) and the alga Chlamydomonas reinhardtii (CrDapL) have been previously reported and it was found that AtDapL and LiDapL showed the same trend of inhibition for different inhibitors in study. It clearly suggests that inhibitors effective against one species may have potential of being active for other species as well ( McKinnie et al, 2014 ).…”

Central IGF‐I Receptors in the Brain are Instrumental to Neuroprotection by Systemically Injected IGF‐I in a Rat Model for Ischemic Stroke

“…The perimembrane domain of megaline is instrumental to the internalization of IGF‐I, and interestingly, the C‐terminal region associates with a glycogen synthase kinase‐3 (GSK‐3), which acts as a key regulator of IGF‐I transport. Mutations of the regulatory site of GSK‐3 modulate IGF‐I internalization, and pharmacological inhibition of GSK‐3 at the choroid plexus increases the internalization of IGF‐I . It would be interesting to test whether GSK‐3 inhibitors can stimulate the transport of systemically injected IGF‐I across the BBB and increase the treatment efficacy of IGF‐I.…”