Citrate sensing by the C4-dicarboxylate/citrate sensor kinase DcuS of Escherichia coli: binding site and conversion of DcuS to a C4-dicarboxylate- or citrate-specific sensor.
The histidine protein kinase DcuS of Escherichia coli senses C(4)-dicarboxylates and citrate by a periplasmic domain. The closely related sensor kinase CitA binds citrate, but no C(4)-dicarboxylates, by a homologous periplasmic domain. CitA is known to bind the three carboxylate and the hydroxyl groups of citrate by sites C1, C2, C3, and H. DcuS requires the same sites for C(4)-dicarboxylate sensing, but only C2 and C3 are highly conserved. It is shown here that sensing of citrate by DcuS required the same sites. Binding of citrate to DcuS, therefore, was similar to binding of C(4)-dicarboxylates but different from that of citrate binding in CitA. DcuS could be converted to a C(4)-dicarboxylate-specific sensor (DcuS(DC)) by mutating residues of sites C1 and C3 or of some DcuS-subtype specific residues. Mutations around site C1 aimed at increasing the size and accessibility of the site converted DcuS to a citrate-specific sensor (DcuS(Cit)). DcuS(DC) and DcuS(Cit) had complementary effector specificities and responded either to C(4)-dicarboxylates or to citrate and mesaconate. The results imply that DcuS binds citrate (similar to the C(4)-dicarboxylates) via the C(4)-dicarboxylate part of the molecule. Sites C2 and C3 are essential for binding of two carboxylic groups of citrate or of C(4)-dicarboxylates; sites C1 and H are required for other essential purposes.
Krämer
J
J
Institut für Mikrobiologie und Weinforschung, Johannes Gutenberg-Universität Mainz, Becherweg 15, 55099 Mainz, Germany.
Fischer
J D
JD
Zientz
E
E
Vijayan
V
V
Griesinger
C
C
Lupas
A
A
Unden
G
G
D000595:Amino Acid Sequence; D001665:Binding Sites; D019343:Citric Acid / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D016000:Cluster Analysis; D019295:Computational Biology; D003998:Dicarboxylic Acids / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005650:Fumarates / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding / Q000187:drug effects; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D014233:Tricarboxylic Acids / Q000378:metabolism / Q000494:pharmacology
D003998:Dicarboxylic Acids; D029968:Escherichia coli Proteins; D005650:Fumarates; D014233:Tricarboxylic Acids; D019343:Citric Acid; D011494:Protein Kinases; C460095:CitA protein, E coli; C469359:DcuS protein, E coli; C028021:tricarballylic acid