Antibiotic resistance and inhibition mechanism of novel aminoglycoside phosphotransferase APH(5) from B. subtilis subsp. subtilis strain RK

Abstract

Bacterial resistance towards aminoglycoside antibiotics mainly occurs because of aminoglycoside phosphotransferases (APHs). It is thus necessary to provide a rationale for focusing inhibitor development against APHs. The nucleotide triphosphate (NTP) binding site of eukaryotic protein kinases (ePKs) is structurally conserved with APHs. However, ePK inhibitors cannot be used against APHs due to cross reactivity. Thus, understanding bacterial resistance at the atomic level could be useful to design new inhibitors against such resistant pathogens. Hence, we carried out in vitro studies of APH from newly deposited multidrug-resistant organism Bacillus subtilis subsp. subtilis strain RK. Enzymatic modification studies of different aminoglycoside antibiotics along with purification and characterization revealed a novel class of APH, i.e., APH(5), with molecular weight 27 kDa approximately. Biochemical analysis of virtually screened inhibitor ZINC71575479 by coupled spectrophotometric assay showed complete enzymatic inhibition of purified APH(5). In silico toxicity study comparison of ZINC71575479 with known inhibitor of APH, i.e., tyrphostin AG1478, predicted its acceptable values for 96 h fathead minnow LC 50 , 48 h Tetrahymena pyriformis IGC 50 , oral rat LD 50 , and developmental toxicity using different QSAR methodologies. Thus, the present study gives novel insight into the aminoglycoside resistance and inhibition mechanism of APH(5) by applying experimental and computational techniques synergistically.

Bacillus subtilisimpact on plant growth, soil health and environment: Dr. Jekyll and Mr. Hyde

“…Recently, B. subtilis subsp. subtilis strain RK, with a novel class of aminoglycoside transferase (AHP5), responsible for resistance against aminoglycoside‐based antibiotics has been identified (Parulekar et al, 2019). The cell wall transporters present in bacteria are crucial for their resistance.…”

Prevalence of Antimicrobial Resistance and Respective Genes among Bacillus spp., a Versatile Bio-Fungicide

“…The absence of certain genes in drug-resistant Bacillus spp. might be because of the presence of some novel mechanisms of resistance, such as enzymatic modifications by aminoglycoside 5-phosphotransferase (aph 5) and methylation in 23S rRNA [ 37 , 38 ]. The phenotypic expression of a certain antimicrobial agents does not always depend upon the resistance genotype and vice versa; that is why the Pearson correlation depicted between resistance phenotypes and genotypes was heterologous.…”

Restoring susceptibility to aminoglycosides: identifying small molecule inhibitors of enzymatic inactivation

“…152 In another study, molecular docking identified an inhibitor of a Bacillus phosphotransferase. 153,154 While these studies are promising, they were limited to in vitro analysis, and further work, including molecular optimization and in vivo experiments, will be required.…”