TY - JOUR

T1 - Identification of additional mechanistically important residues in the multidrug transporter styMdtM of Salmonella Typhi

AU - Shaheen, Aqsa

AU - Tariq, Anam

AU - Ismat, Fouzia

AU - Naveed, Hammad

AU - De Zorzi, Rita

AU - Iqbal, Mazhar

AU - Storici, Paola

AU - Mirza, Osman

AU - Walz, Thomas

AU - Rahman, Moazur

N1 - Publisher Copyright: © 2023 Informa UK Limited, trading as Taylor & Francis Group.

PY - 2023

Y1 - 2023

N2 - Multidrug efflux is a well-established mechanism of drug resistance in bacterial pathogens like Salmonella Typhi. styMdtM (locus name; STY4874) is a multidrug efflux transporter of the major facilitator superfamily expressed in S. Typhi. Functional assays identified several residues important for its transport activity. Here, we used an AlphaFold model to identify additional residues for analysis by mutagenesis. Mutation of peripheral residue Cys185 had no effect on the structure or function of the transporter. However, substitution of channel-lining residues Tyr29 and Tyr231 completely abolished transport function. Finally, mutation of Gln294, which faces peripheral helices of the transporter, resulted in the loss of transport of some substrates. Crystallization studies yielded diffraction data for the wild-type protein at 4.5 Å resolution and allowed the unit cell parameters to be established as a = b = 64.3 Å, c = 245.4 Å, α = β = γ = 90°, in space group P4. Our studies represent a further stepping stone towards a mechanistic understanding of the clinically important multidrug transporter styMdtM. Communicated by Ramaswamy H. Sarma.

AB - Multidrug efflux is a well-established mechanism of drug resistance in bacterial pathogens like Salmonella Typhi. styMdtM (locus name; STY4874) is a multidrug efflux transporter of the major facilitator superfamily expressed in S. Typhi. Functional assays identified several residues important for its transport activity. Here, we used an AlphaFold model to identify additional residues for analysis by mutagenesis. Mutation of peripheral residue Cys185 had no effect on the structure or function of the transporter. However, substitution of channel-lining residues Tyr29 and Tyr231 completely abolished transport function. Finally, mutation of Gln294, which faces peripheral helices of the transporter, resulted in the loss of transport of some substrates. Crystallization studies yielded diffraction data for the wild-type protein at 4.5 Å resolution and allowed the unit cell parameters to be established as a = b = 64.3 Å, c = 245.4 Å, α = β = γ = 90°, in space group P4. Our studies represent a further stepping stone towards a mechanistic understanding of the clinically important multidrug transporter styMdtM. Communicated by Ramaswamy H. Sarma.

KW - E. coli MdfA

KW - efflux pump

KW - MdtM

KW - multidrug resistance

KW - Salmonella Typhi

KW - site-directed mutagenesis

U2 - 10.1080/07391102.2023.2263882

DO - 10.1080/07391102.2023.2263882

M3 - Journal article

C2 - 37787617

AN - SCOPUS:85173510897

JO - Journal of Biomolecular Structure and Dynamics

JF - Journal of Biomolecular Structure and Dynamics

SN - 0739-1102

ER -