Unravelling the Structural and Molecular Basis Responsible for the Anti-Biofilm Activity of Zosteric Acid

Abstract

The natural compound zosteric acid, or p-(sulfoxy)cinnamic acid (ZA), is proposed as an alternative biocide-free agent suitable for preventive or integrative anti-biofilm approaches. Despite its potential, the lack of information concerning the structural and molecular mechanism of action involved in its anti-biofilm activity has limited efforts to generate more potent anti-biofilm strategies. In this study a 43-member library of small molecules based on ZA scaffold diversity was designed and screened against Escherichia coli to understand the structural requirements necessary for biofilm inhibition at sub-lethal concentrations. Considerations concerning the relationship between structure and anti-biofilm activity revealed that i) the para-sulfoxy ester group is not needed to exploit the anti-biofilm activity of the molecule, it is the cinnamic acid scaffold that is responsible for anti-biofilm performance; ii) the anti-biofilm activity of ZA derivatives depends on the presence of a carboxylate anion and, consequently, on its hydrogen-donating ability; iii) the conjugated aromatic system is instrumental to the anti-biofilm activities of ZA and its analogues. Using a protein pull-down approach, combined with mass spectrometry, the herein-defined active structure of ZA was matrix-immobilized, and was proved to interact with the E. coli NADH:quinone reductase, WrbA, suggesting a possible role of this protein in the biofilm formation process.

Low density polyethylene functionalized with antibiofilm compounds inhibits Escherichia coli cell adhesion

“…5a,b) when they are free in solution, showing emission maxima at 395 and 443 nm for p-aminosalicylic acid (k exc 5 325 nm) and p-aminocinnamic acid (k exc 5 380 nm), respectively. 16 This property allowed us to evaluate the fluorescence of the solid surface of the functionalized polyethylene.…”

“…Indeed, cells use ROS as a signal or cue to adapt to a changing environment. 46 Recently, it has been found that E. coli proteins targeted by ZA 16 and SA 23 are widespread with a high percentage of identity in a wide range of microorganisms, included several E. coli pathogenic stains and bacteria involved in serious disease. Thus, it is possible that LDPE-CA and LDPE-ZA could exert their activity also against other bacterial strains.…”

“…10 Therefore, new approaches become imperative, and a serious change in our prospective is necessary: instead of fighting biofilm with antimicrobial materials and coatings, the efforts should be directed toward developing innovative anti-biofilm materials with functional features targeting molecular determinants of biofilm genesis (e.g., substratum adherence), disarming microorganisms without killing them. 15,16 Active substances with well-known anti-biofilm activity at low concentrations would not be leached from the surface, avoiding the problem of the compound kinetics release, and providing long-term protection against microbial colonization. Depriving microorganisms of their biofilm-specific traits, without affecting their existence, may also decrease selection pressure for drug-resistant mutations, restoring the efficacy of the current arsenal of antimicrobial agents.…”

“…10,20 However, to the best of our knowledge, nobody investigated the possibility to design a non-leaching, long lasting, antibiofilm material to prevent colonization of polymeric materials, by using biocide-free compounds able to hinder biofilm formation. In our previous study, 16 we designed and screened against Escherichia coli a 43-member library of small molecules based on zosteric acid scaffold diversity to understand the structural requirements necessary for biofilm inhibition, and to identify functional groups that could be exploited for the covalent linkage to an abiotic surface. This work revealed that p-aminocinnamic acid, targeting WrbA a FMN-dependent oxido-reductase, 16 is able to significantly reduce E. coli biofilm formation, and it could be considered as an excellent candidate to be covalently linked to a polymeric support due to the presence of an amine at the para position on the phenyl ring.…”

“…In our previous study, 16 we designed and screened against Escherichia coli a 43-member library of small molecules based on zosteric acid scaffold diversity to understand the structural requirements necessary for biofilm inhibition, and to identify functional groups that could be exploited for the covalent linkage to an abiotic surface. This work revealed that p-aminocinnamic acid, targeting WrbA a FMN-dependent oxido-reductase, 16 is able to significantly reduce E. coli biofilm formation, and it could be considered as an excellent candidate to be covalently linked to a polymeric support due to the presence of an amine at the para position on the phenyl ring. Besides cinnamic acid analogues, another class of derivatives related to the scaffold of salicylic acid has been investigated.…”