TY  - JOUR
A1  - Heuveling, Johanna
A1  - Frochaux, Violette
A1  - Ziomkowska, Joanna
A1  - Wawrzinek, Robert
A1  - Wessig, Pablo
A1  - Herrmann, Andreas
A1  - Schneider, Erwin
T1  - Conformational changes of the bacterial type I ATP-binding cassette importer HisQMP(2) at distinct steps of the catalytic cycle
JF  - Biochimica et biophysica acta : Biomembranes
N2  - Prokaryotic solute binding protein-dependent ATP-binding cassette import systems are divided into type land type II and mechanistic differences in the transport process going along with this classification are under intensive investigation. Little is known about the conformational dynamics during the catalytic cycle especially concerning the transmembrane domains. The type I transporter for positively charged amino acids from Salmonella enterica serovar Typhimurium (1A0-Hi5QMP2) was studied by limited proteolysis in detergent solution in the absence and presence of co-factors including ATP, ADP, LAO/arginine, and Mg2+ ions. Stable peptide fragments could be obtained and differentially susceptible cleavage sites were determined by mass spectrometry as Lys-258 in the nucleotide-binding subunit, HisP, and Arg-217/Arg-218 in the transmembrane subunit, HisQ In contrast, transmembrane subunit HisM was gradually degraded but no stable fragment could be detected. HisP and HisQ were equally resistant under pre- and post-hydrolysis conditions in the presence of arginine-loaded solute-binding protein LAO and ATP/ADP. Some protection was also observed with LAO/arginine alone, thus reflecting binding to the transporter in the apo-state and transmembrane signaling. Comparable digestion patterns were obtained with the transporter reconstituted into proteoliposomes and nanodiscs. Fluorescence lifetime spectroscopy confirmed the change of HisQ(R218) to a more apolar microenvironment upon ATP binding and hydrolysis. Limited proteolysis was subsequently used as a tool to study the consequences of mutations on the transport cycle. Together, our data suggest similar conformational changes during the transport cycle as described for the maltose ABC transporter of Escherichia coli, despite distinct structural differences between both systems.
KW  - ABC transporter
KW  - Type I importer
KW  - Histidine transport
KW  - Limited proteolysis
KW  - Fluorescence lifetime
KW  - Altemate access model
Y1  - 2014
U6  - https://doi.org/10.1016/j.bbamem.2013.08.024
SN  - 0005-2736
SN  - 0006-3002
VL  - 1838
IS  - 1
SP  - 106
EP  - 116
PB  - Elsevier
CY  - Amsterdam
ER  -