Comparative Analysis of Mesophilic YqfB-Type Amidohydrolases

Abstract

The widespread superfamily of the human activating signal cointegrator homology (ASCH) domain was identified almost 20 years ago; however, the amount of experimental data regarding the biological function of the domain is scarce. With this study, we aimed to determine the putative cellular functions of four hypothetical ASCH domain-containing amidohydrolase YqfB analogues by investigating their activity towards various N-acylated cytosine derivatives, including potential nucleoside-derived prodrugs, as well as their ability to bind/degrade nucleic acids in vitro. According to determined kinetic parameters, N4-acetylcytidine is assumed to be the primary substrate for amidohydrolases. Despite the similarity to the proteins containing the PUA domain, no nucleic acid binding activity was detected for YqfB-like proteins, suggesting that, in vivo, these enzymes are a part of the pyrimidine salvage pathway. We also demonstrate the possibility of the expression of YqfB-type amidohydrolases in both prokaryotic and eukaryotic hosts. The small protein size and remarkable halotolerance of YqfB-type amidohydrolases are of great interest for further fundamental research and biotechnological applications

Bacterial amidohydrolases and modified 5-fluorocytidine compounds: Novel enzyme-prodrug pairs

“…Moreover, the halotolerance of these enzymes opens up the possibility of delivering them into the tumor microenvironment as a protein that is likely to remain stable and active even in biological fluids such as blood. In a recent study, analogues of YqfB amidohydrolase found in other microorganisms showed better catalytic efficiency compared to YqfB for the primary substrate N 4 -acetylcytidine [ 17 ]. This suggests that more studies may lead to even better amidohydrolase enzyme-prodrug combinations to achieve the highest therapeutic efficacy.…”

“…5-fluorocytidine was purchased from Biosynth Carbosynth (Berkshire, UK). The synthesis of 5-fluoro- N 4 -benzoylcytidine and 5-fluoro- N 4 -pivaloylcytidine was described previously [ 17 ]. The synthesis of 5-fluoro- N 4 -acetylcytidine and 5-fluoro- N 4 -[3-indolepropionyl]cytidine (Fig A in S1 File ) is described in the Supporting information.…”

“…5-fluorocytidine was purchased from Biosynth Carbosynth (Berkshire, UK). The synthesis of 5-fluoro-N 4 -benzoylcytidine and 5-fluoro-N 4 -pivaloylcytidine was described previously [17]. The synthesis of 5-fluoro-N 4…”

“…Previously, the YqfB [15] and D8_RL [16] proteins were described as small amidohydrolases (103 aa and 128 aa respectively) active towards N 4 -acylated cytidines, e.g., N 4 -acetylcytidine [15][16][17]. YqfB is the product of the yqfB gene in Escherichia coli, while D8_RL was isolated from metagenomic libraries.…”