AiiA, an enzyme that inactivates the acylhomoserine lactone quorum-sensing signal and attenuates the virulence of Erwinia carotovora

Abstract

N-acylhomoserine lactones, known as autoinducers (AIs), are widely conserved signal molecules present in quorum-sensing systems of many Gram-negative bacteria. AIs are involved in the regulation of diverse biological functions, including expression of pathogenic genes in the plant pathogens Pseudomonas solanacearum, several Erwinia species, and the human pathogen Pseudomonas aeruginosa. A bacterial isolate, Bacillus sp. 240B1, is capable of enzymatic inactivation of AIs. The gene (aiiA) for AI inactivation from Bacillus sp. 240B1 has been cloned and shown to encode a protein of 250 amino acids. Sequence alignment indicates that AiiA contains a ''HXHXDH'' zinc-binding motif that is conserved in several groups of metallohydrolases. Site-directed mutagenesis showed that conserved aspartate and most histidine residues are required for AiiA activity. Expression of aiiA in transformed Erwinia carotovora strain SCG1 significantly reduces the release of AI, decreases extracellular pectolytic enzyme activities, and attenuates pathogenicity on potato, eggplant, Chinese cabbage, carrot, celery, cauliflower, and tobacco. Our results indicate that the AI-inactivation approach represents a promising strategy for prevention of diseases in which virulence is regulated by AIs.

Exploiting Quorum Sensing To Confuse Bacterial Pathogens

“…Although the aiiA gene did not share significant homology with any known sequences, it did encode an HXHXD sequence motif common to metallo-␤-lactamases. Purified AiiA protein inactivated multiple AHLs in vitro (57), and biochemical analysis confirmed that AiiA functioned as a lactonase (85). Importantly, when cloned into the plant pathogen Erwinia carotovora, the aiiA gene reduced AHL release from the pathogen, decreased QS-controlled enzyme activities, and reduced soft rot disease symptoms on potato, eggplant, Chinese cabbage, carrot, and celery plants (57).…”

“…AHL lactonases are metalloproteins (with the exception of AiiM) that hydrolyze the ester bond of the homoserine lactone ring to yield the corresponding acyl homoserine molecule (Fig. 3) (57,85). Hydrolysis of this ester bond can also occur spontaneously at alkaline pH, and acidic conditions can restore the bond and AHL signaling activity following biological or abiotic hydrolysis (86).…”

“…The first report of biological inactivation of AHL signaling molecules was published in 2000 following the screening of more than 500 bacterial field isolates and lab strains. An AHLinactivating isolate was characterized to be a Bacillus sp., and genetic studies allowed for the identification of the gene responsible for the inactivation activity, which was designated aiiA (autoinducer inactivation gene) (57). Although the aiiA gene did not share significant homology with any known sequences, it did encode an HXHXD sequence motif common to metallo-␤-lactamases.…”

“…Purified AiiA protein inactivated multiple AHLs in vitro (57), and biochemical analysis confirmed that AiiA functioned as a lactonase (85). Importantly, when cloned into the plant pathogen Erwinia carotovora, the aiiA gene reduced AHL release from the pathogen, decreased QS-controlled enzyme activities, and reduced soft rot disease symptoms on potato, eggplant, Chinese cabbage, carrot, and celery plants (57). Additionally, transgenic potato and tobacco plants carrying the aiiA gene showed enhanced resistance to E. carotovora infection (Fig.…”

In Vitro Characterization of a Recombinant AHL-Lactonase from Bacillus cereus Isolated from a Striped Catfish (Pangasianodon hypophthalmus) Pond

“…and other species [9,[12][13][14]. Genetically-modified Erwinia carotovora and Pseudomonas aeruginosa expressing AHL-lactonase showed decreased production of virulence factors and attenuated virulence [15,16]. AHL-lactonase has been over-expressed successfully in E. coli and Pichia pastoris expression system as a soluble heterologous protein [17,18].…”