Short-chain diamines are the physiological substrates of PACE family efflux pumps

Abstract

Acinetobacter baumannii has rapidly emerged as a major cause of gram-negative hospital infections worldwide. A. baumannii encodes for the transport protein AceI, which confers resistance to chlorhexidine, a widely used antiseptic. AceI is also the prototype for the recently discovered proteobacterial antimicrobial compound efflux (PACE) family of transport proteins that confer resistance to a range of antibiotics and antiseptics in many gram-negative bacteria, including pathogens. The gene encoding AceI is conserved in the core genome of A. baumannii, suggesting that it has an important primordial function. This is incongruous with the sole characterized substrate of AceI, chlorhexidine, an entirely synthetic biocide produced only during the last century. Here we investigated a potential primordial function of AceI and other members of the PACE family in the transport of naturally occurring polyamines. Polyamines are abundant in living cells, where they have physiologically important functions and play multifaceted roles in bacterial infection. Gene expression studies revealed that the aceI gene is induced in A. baumannii by the short-chain diamines cadaverine and putrescine. Membrane transport experiments conducted in whole cells of A. baumannii and Escherichia coli and also in proteoliposomes showed that AceI mediates the efflux of these short-chain diamines when energized by an electrochemical gradient. Assays conducted using 8 additional diverse PACE family proteins identified 3 that also catalyze cadaverine transport. Taken together, these results demonstrate that short-chain diamines are common substrates for the PACE family of transport proteins, adding to their broad significance as a novel family of efflux pumps.

Spermidine and spermine are the natural substrates of theAcinetobacter baumanniiAmvA multidrug efflux pump

“…Whether produced endogenously or present in the environment, polyamines can be toxic in high amounts. Several efflux systems have been reported to facilitate polyamine transport, such as members of the small multidrug resistance (SMR) (20), major facilitator superfamily (MFS) (21), and PACE families (11).…”

Assembly and regulation of the chlorhexidine-specific efflux pump AceI

“…Studies of AceI homologs from other species have expanded the substrates for PACE family proteins to include synthetic biocides. These molecules include benzalkonium, acriflavine, proflavine, and dequalinium in addition to chlorhexidine (8) and, more recently, naturally occurring short-chain diamines (9). Transport experiments conducted in whole cells and proteoliposomes suggest that the prototype protein AceI use electrochemical proton gradient to translocate substrates (6,9).…”

“…These molecules include benzalkonium, acriflavine, proflavine, and dequalinium in addition to chlorhexidine (8) and, more recently, naturally occurring short-chain diamines (9). Transport experiments conducted in whole cells and proteoliposomes suggest that the prototype protein AceI use electrochemical proton gradient to translocate substrates (6,9). AceI shares a similar size and predicted secondary structure as the SMR family, to date but no structural or functional similarities have been described for these two families.…”

“…Studies of AceI homologs from other species have expanded the substrates for PACE family proteins to include synthetic biocides. These molecules include benzalkonium, acriflavine, proflavine, and dequalinium in addition to chlorhexidine ( 8 ) and, more recently, naturally occurring short-chain diamines ( 9 ).…”

“…Transport experiments conducted in whole cells and proteoliposomes suggest that the prototype protein AceI use electrochemical proton gradient to translocate substrates ( 6 , 9 ). AceI shares a similar size and predicted secondary structure as the SMR family, to date but no structural or functional similarities have been described for these two families.…”