Engineering Hybrid Chemotaxis Receptors in Bacteria
Supporting Information for
Engineering Hybrid Chemotaxis Receptors in Bacteria Shuangyu Bi1, Abiola M. Pollard1, Yiling Yang1, Fan Jin1, and Victor Sourjik*
Max Planck Institute for Terrestrial Microbiology & LOEWE Center for Synthetic Microbiology (SYNMIKRO), Marburg, Germany
*
To whom correspondence should be addressed:
Victor Sourjik, Ph.D. Max Planck Institute for Terrestrial Microbiology & LOEWE Center for Synthetic Microbiology (SYNMIKRO) D-35043 Marburg, Germany Tel. (+49) 6421 28 21400; Fax (+49) 6421 28 21485 E-mail [email protected]
1
Supplementary Figures
Figure S1. Expression of hybrid receptors. (a) The expression of hybrid receptors in E. coli cheR cheB strain VH1 was tested using immunoblotting. VH1 expressing the wild-type Tar and VH1 carrying the empty vector (pKG116) were used as controls. (b) Relative expression levels of the hybrid receptors, quantified using ImageJ. The expression level of the wild-type Tar was normalized to be one. Error bars indicate the standard errors of two biological replicates.
2
Figure S2. FRET responses of NarQ-Tar and Tar to nitrite and nitrate. (a) FRET measurements of buffer-adapted E. coli cells expressing NarQ-Tar as the sole receptor responding to a stepwise addition (down arrow) and subsequent removal (up arrow) of the indicated concentrations of NaNO2. (b) FRET measurements of the response of the wild-type Tar to NaNO3 or NaNO2 as indicated. The response to D,L--methyl-aspartate (MeAsp) was measured as a control.
3
4
Figure S3. FRET responses of McpS-Tar and Tar to succinate, citrate or L-malate. (a, b) FRET measurements and dose-response curves of buffer-adapted E. coli cells expressing McpS-Tar as the sole receptor to the indicated concentrations of succinate (a) or citrate (b). (c) FRET measurements of the wild-type Tar response to L-malate, succinate, and citrate. (d) FRET measurements of the response of McpS-Tar and wild-type Tar to acetate.
5
Figure S4. FRET measurement of the ECA0434-Tar response to NaNO2. Measurement was performed as in Figure 3d.
6
Figure S5. Responses of CitA-Tar, Tar, and Tar°-T303I to citrate, isocitrate, itaconic and maleic acids. (a) The responses of the hybrid Tar43CitA55Tar184 and Tar to citrate measured by an agar plate assay. The site of the ligand source and inoculated cells are shown by a red dotted 7
line and by a yellow dot, respectively. Relative spreading of cells toward or away from the ligand source is indicated. (b) FRET measurements and dose response of buffer-adapted E. coli cells expressing Tar43CitA55Tar184 as the sole receptor to the indicated concentrations of isocitrate. (c) FRET measurements of the response of buffer-adapted E. coli cells expressing wild-type Tar to a stepwise addition (down arrow) and subsequent removal (up arrow) of the indicated concentrations of isocitrate, itaconic and maleic acids. (d-f) FRET measurements of the response of Tar43CitA55Tar184 (d), Tar°-T303I (e), and receptorless E. coli (f) to itaconic and maleic acids.
8
Figure S6. FRET measurements of the response of Tar and Tar°-T303I to L-proline.
9
Figure S7. Approach used for the selection of functional McpC-Tar hybrids from the McpC[1-350]-NNNNN-Tar[272-553] library.
10
Figure S8. FRET responses of McpC350Tar267 to newly identified ligands. Measurements were performed for buffer-adapted E. coli cells expressing McpC350Tar267 as the sole receptor responding to a stepwise addition (down arrow) and subsequent removal (up arrow) of the indicated concentrations of ligands.
11
12
Figure S9. Responses of wild-type Tar or Tar°-T303I to novel ligands for McpC, McpG or McpS. (a) The values of the fluorescence intensities in the analysis regions emitted by E. coli cells expressing the wild-type Tar or Tar°-T303I responding to the novel ligands of McpC, McpG, or McpS in microfluidic assays. The measurements were done at 30 min for 300 M or 1 mM of ligands, as indicated. (b) FRET responses of buffer-adapted E. coli cells expressing the wild-type Tar to the indicated ligands.
13
Figure S10. FRET responses of McpG348Tar268 to the indicated ligands.
14
Figure S11. FRET responses of McpS295Tar200 to the indicated ligands.
15
Supplementary Tables
Table S1. Plasmids and strains used in this study Genotype or phenotype
Induction
Source or reference
Plasmids pKG116
Expression vector, Camr
-
1
pHP2
Tar expression plasmid with added NotI and deleted
-
2
2 µM sodium
3
NdeI restriction site, Ampr pPA791
Tar∆(44-183)/T303I expression plasmid, Camr
salicylate pVS1092
Tar expression plasmid, Camr
2 µM sodium
4
salicylate pVS88
CheY-EYFP / CheZ-ECFP expression plasmid, Ampr
50 µM IPTG
5
pOB30
GFP expression plasmid, Ampr
50 µM IPTG
Gift of Olga Besharova
pSB1
pSB2
pSB3
pSB4
pSB5
pSB6
Hybrid Tar[1-43]-McpC[49-270]-Tar[184-553]
2 µM sodium
expression plasmid, pKG116 derivative
salicylate
Hybrid McpC[1-286]-Tar[200-553] expression plasmid,
2 µM sodium
pKG116 derivative
salicylate
Hybrid McpC[1-350]-Tar[267-553] expression plasmid,
2 µM sodium
pKG116 derivative
salicylate
Hybrid McpC[1-340] and Tar[257-553] expression
2 µM sodium
plasmid, pKG116 derivative
salicylate
Hybrid McpC[1-472]-Tar[361-417]-McpC[530-655]
2 µM sodium
expression plasmid, pKG116 derivative
salicylate
Hybrid McpC[1-350]-YSYEH-Tar[272-553] expression
2 µM sodium
plasmid, pKG116 derivative
salicylate 16
This study
This study
This study
This study
This study
This study
pSB7
pSB8
pSB9
pSB10
pSB11
pSB12
Hybrid Tar[1-43]-CitA[55-173]-Tar[184-553]
2 µM sodium
expression plasmid, pKG116 derivative
salicylate
Hybrid CitA[1-193]-Tar[204-553] expression plasmid,
2 µM sodium
pKG116 derivative
salicylate
Hybrid ECA0434[1-385]-Tar[267-553] expression
2 µM sodium
plasmid, pKG116 derivative
salicylate
Hybrid McpS[1-295]-Tar[200-553] expression plasmid,
0.7 µM sodium
pKG116 derivative
salicylate
Hybrid NarQ[1-217]-Tar[257-553] expression plasmid,
2 µM sodium
pKG116 derivative
salicylate
Hybrid McpG[1-348]-Tar[268-553] expression plasmid,
0.8 µM sodium
pKG116 derivative
salicylate
This study
This study
This study
This study
This study
6
Strains UU1250
E. coli RP437 ΔaerΔtsrΔ(tar-tap)Δtrg
-
7
VS181
E. coli RP437 Δ(cheYcheZ)ΔaerΔtsrΔ(tar-tap)Δtrg
-
5
VH1
E. coli RP437
-
8
Δ(cheRcheB)Δ(cheYcheZ)ΔaerΔtsrΔ(tar-tap)Δtrg
17
Table S2. List of all compounds screened for McpC350Tar267
Name
Response of hybrid receptor
Putrescine
Nb
N-phthaloyl-Lglutamic acid N-acetyl-L-glutamic acid Glycyl-L-aspartic acid D,L-lactamide Glycyl-Lmethionine
Response of Tara
Name D-2-phospho-glyceric acid D-3-phospho-glyceric acid
N N
O-phospho-D-serine
N N N
O-phospho-Lthreonine O-phospho-L-tyrosine O-phospho-Dtyrosine O-phospho-Dthreonine O-phospho-L-serine m-Hydroxy-phenyl acetic acid L-methionine sulfoxide Hydroxylamine Methylamine Parabanic acid
Response of hybrid receptor N N N N N N
Adenine
N
L-ornithine
N
L-cysteinyl-glycine
N
Ala-Asp
N
L-homoserine L-citrulline Ala-Gly
-c N N
-Hydroxy-butyric acid
N
n-Butylamine
N
-Hydroxy-butyric acid
N
Ethylamine
N
N N
Ethanolamine Ethylenediamine
N N
N
Glucuronamide
N
N
Agmatine
N
Histamine
N
-Phenylethyl-amine
N
Tyramine
N
-Keto-valeric acid Ala-Gln 3-Hydroxy-2butanone -Amino-n-caproica cid -Amino-n-valeric acid p-Hydroxy-phenyl acetic acid D,L--amino-nbutyric acid
-
N
N
N -
N 18
N N N N N N N
Response of Tara
-Amino-n-butyric acid
N
Acetamide
N
-Amino-n-valeric acid
N
Formamide
N
D-alanine
-
D-aspartic acid D-methionine D-asparagine D-threonine D-cysteine D-glutamic acid D-lysine D-valine D-serine Inosine Xanthine Uridine Acetamide D,L-carnitine D,L-octopamine 2,3-Butanediol N-acetyl-L-cysteine
N N N N N N N N N N N N N N N N N
-Hydroxy-glutaric acid--lactone
N
a
D,L--amino-caprylic acid Adenosine Cytidine Cytosine Guanine Guanosine Thymine Thymidine Uracil Allantoin Alloxan Uric acid Xanthosine Ala-Glu Sec-butylamine Dihydroxyaceton 2,3-Butanone S-methyl-L-cysteine
N
-Hydroxy-butyric acid
N N N N N N N N N N N N N N N N N N N
Measured only for compounds that induce a chemotactic response via the hybrid receptor. bNo
chemotactic response. cRepellent response.
19
Table S3. List of all compounds screened for McpG348Tar268
Name
Response of hybrid receptor
L-aspartic acid
Nb
L-glutamic acid
N
L-alanine L-asparagine L-cysteine L-arginine L-glutamine Glycine L-histidine L-isoleucine L-leucine L-lysine L-methionine
+c N + N N N N N N N N
L-phenylalanine
N
L-proline L-serine L-threonine L-valine L-tryptophan L-tyrosine L-homoserine L-ornithine D-alanine D-aspartic acid D-methionine D-asparagine D-threonine D-cysteine D-glutamic acid D-lysine D-valine D-serine N-acetyl-Lglutamic acid L-citrulline
+ N N N N N + N N N N N N N N N N N
Response of Tara
+ +
Name Urea N-phthaloyl-Lglutamic acid L-pyroglutamic acid Hydroxylamine Methylamine N-amylamine N-butylamine Ethylamine Ethanolamine Ethylenediamine Putrescine Agmatine Histamine -Phenylethylamine Tyramine Acetamide Formamide Adenine Adenosine Cytidine Cytosine Guanine Guanosine Thymine Thymidine Uracil Uridine Inosine Xanthine Xanthosine Uric acid Alloxan
N
N
Response of hybrid receptor N N N N N N N N N + + + N N N N N N N N N N N N N N N N N N N N
N
Allantoin
N
N
Parabanic acid
N
20
Response of Tara
N N N
D,L--amino-nbutyric acid D,L--aminocaprylic acid Glucuronamide a
-Amino-n-caproic acid
N
-Amino-n-valeric acid D,L-lactamide
N N
N N N
Measured only for compounds that induce a chemotactic response via the hybrid receptor. bNo
chemotactic response. cAttractant response.
21
Table S4. List of all compounds screened for McpS295Tar200
Name
Response of hybrid receptor
Name
Response of hybrid receptor
L-aspartic acid L-glutamic acid
+b Nc
D-asparagine D-glutamic acid
N N
L-alanine
N
D,L--amino-nbutyric acid
N
L-asparagine
N
-Amino-n-butyric acid
N
L-cysteine
N
-Amino-n-caproica cid
N
L-arginine
N
-Amino-valeric acid
N
L-glutamine
N
Glycine L-histidine
N N
L-isoleucine
N
-Hydroxy-butyric acid
N
L-leucine
N
-Hydroxy-butyric acid
N
L-lysine
N
L-methionine
N
L-phenylalanine L-proline L-serine L-threonine
N N N N
L-valine
N
L-tryptophan
N
L-tyrosine
N
Methyl-pyruvate
N
Oxaloacetic acid
N
D,L--hydroxybutyric acid D-aspartic acid Itaconic acid
Response of Tara +
-Amino-n-valeric acid Propionic acid Caproic acid
-Keto-valeric acid -Hydroxy-butyric acid Citraconic acid L-tartaric acid Quinolinic acid Nicotinic acid -Amino-levulinic acid D,L-mevalonic acid
N N +
N N N N N N N
N
N
Methyl-pyruvate
N
N +
Oxaloacetic acid Valeric acid
N +
22
N
N
-Hydroxy-glutaric acid--lactone Acetoacetic acid Mono methylsuccinate
N
Response of Tara
+
N
N
N
-Keto-butyric acid
N
D-tartaric acid
+
N
a
Measured only for compounds that induce chemotactic response via the hybrid receptor.
b
Attractant response. cNo chemotactic response.
23
Supplementary References 1. Buron-Barral, M. C., Gosink, K. K., and Parkinson, J. S. (2006) Loss- and gain-of-function mutations in the F1-HAMP region of the Escherichia coli aerotaxis transducer Aer. J. Bacteriol. 188, 3477-3486. 2. Pham, H. T., and Parkinson, J. S. (2011) Phenol sensing by Escherichia coli chemoreceptors: a nonclassical mechanism. J. Bacteriol. 193, 6597-6604. 3. Gosink, K. K., Buron-Barral, M. C., and Parkinson, J. S. (2006) Signaling interactions between the aerotaxis transducer Aer and heterologous chemoreceptors in Escherichia coli. J. Bacteriol. 188, 3487-3493. 4. Yang, Y., and Sourjik, V. (2012) Opposite responses by different chemoreceptors set a tunable preference point in Escherichia coli pH taxis. Mol. Microbiol. 86, 1482-1489. 5. Sourjik, V., and Berg, H. C. (2004) Functional interactions between receptors in bacterial chemotaxis. Nature 428, 437-441. 6. Reyes-Darias, J. A., Garcia, V., Rico-Jimenez, M., Corral-Lugo, A., Lesouhaitier, O., Juarez-Hernandez, D., Yang, Y., Bi, S., Feuilloley, M., Munoz-Rojas, J., Sourjik, V., and Krell, T. (2015) Specific gamma-aminobutyrate chemotaxis in pseudomonads with different lifestyle. Mol. Microbiol. 97, 488-501. 7. Ames, P., Studdert, C. A., Reiser, R. H., and Parkinson, J. S. (2002) Collaborative signaling by mixed chemoreceptor teams in Escherichia coli. Proc. Natl. Acad. Sci. U.S.A. 99, 7060-7065. 8. Oleksiuk, O., Jakovljevic, V., Vladimirov, N., Carvalho, R., Paster, E., Ryu, W. S., Meir, Y., Wingreen, N. S., Kollmann, M., and Sourjik, V. (2011) Thermal robustness of signaling in bacterial chemotaxis, Cell 145, 312-321.
24
Engineering Hybrid Chemotaxis Receptors in Bacteria Shuangyu Bi1, Abiola M. Pollard1, Yiling Yang1, Fan Jin1, and Victor Sourjik*
Max Planck Institute for Terrestrial Microbiology & LOEWE Center for Synthetic Microbiology (SYNMIKRO), Marburg, Germany
*
To whom correspondence should be addressed:
Victor Sourjik, Ph.D. Max Planck Institute for Terrestrial Microbiology & LOEWE Center for Synthetic Microbiology (SYNMIKRO) D-35043 Marburg, Germany Tel. (+49) 6421 28 21400; Fax (+49) 6421 28 21485 E-mail [email protected]
1
Supplementary Figures
Figure S1. Expression of hybrid receptors. (a) The expression of hybrid receptors in E. coli cheR cheB strain VH1 was tested using immunoblotting. VH1 expressing the wild-type Tar and VH1 carrying the empty vector (pKG116) were used as controls. (b) Relative expression levels of the hybrid receptors, quantified using ImageJ. The expression level of the wild-type Tar was normalized to be one. Error bars indicate the standard errors of two biological replicates.
2
Figure S2. FRET responses of NarQ-Tar and Tar to nitrite and nitrate. (a) FRET measurements of buffer-adapted E. coli cells expressing NarQ-Tar as the sole receptor responding to a stepwise addition (down arrow) and subsequent removal (up arrow) of the indicated concentrations of NaNO2. (b) FRET measurements of the response of the wild-type Tar to NaNO3 or NaNO2 as indicated. The response to D,L--methyl-aspartate (MeAsp) was measured as a control.
3
4
Figure S3. FRET responses of McpS-Tar and Tar to succinate, citrate or L-malate. (a, b) FRET measurements and dose-response curves of buffer-adapted E. coli cells expressing McpS-Tar as the sole receptor to the indicated concentrations of succinate (a) or citrate (b). (c) FRET measurements of the wild-type Tar response to L-malate, succinate, and citrate. (d) FRET measurements of the response of McpS-Tar and wild-type Tar to acetate.
5
Figure S4. FRET measurement of the ECA0434-Tar response to NaNO2. Measurement was performed as in Figure 3d.
6
Figure S5. Responses of CitA-Tar, Tar, and Tar°-T303I to citrate, isocitrate, itaconic and maleic acids. (a) The responses of the hybrid Tar43CitA55Tar184 and Tar to citrate measured by an agar plate assay. The site of the ligand source and inoculated cells are shown by a red dotted 7
line and by a yellow dot, respectively. Relative spreading of cells toward or away from the ligand source is indicated. (b) FRET measurements and dose response of buffer-adapted E. coli cells expressing Tar43CitA55Tar184 as the sole receptor to the indicated concentrations of isocitrate. (c) FRET measurements of the response of buffer-adapted E. coli cells expressing wild-type Tar to a stepwise addition (down arrow) and subsequent removal (up arrow) of the indicated concentrations of isocitrate, itaconic and maleic acids. (d-f) FRET measurements of the response of Tar43CitA55Tar184 (d), Tar°-T303I (e), and receptorless E. coli (f) to itaconic and maleic acids.
8
Figure S6. FRET measurements of the response of Tar and Tar°-T303I to L-proline.
9
Figure S7. Approach used for the selection of functional McpC-Tar hybrids from the McpC[1-350]-NNNNN-Tar[272-553] library.
10
Figure S8. FRET responses of McpC350Tar267 to newly identified ligands. Measurements were performed for buffer-adapted E. coli cells expressing McpC350Tar267 as the sole receptor responding to a stepwise addition (down arrow) and subsequent removal (up arrow) of the indicated concentrations of ligands.
11
12
Figure S9. Responses of wild-type Tar or Tar°-T303I to novel ligands for McpC, McpG or McpS. (a) The values of the fluorescence intensities in the analysis regions emitted by E. coli cells expressing the wild-type Tar or Tar°-T303I responding to the novel ligands of McpC, McpG, or McpS in microfluidic assays. The measurements were done at 30 min for 300 M or 1 mM of ligands, as indicated. (b) FRET responses of buffer-adapted E. coli cells expressing the wild-type Tar to the indicated ligands.
13
Figure S10. FRET responses of McpG348Tar268 to the indicated ligands.
14
Figure S11. FRET responses of McpS295Tar200 to the indicated ligands.
15
Supplementary Tables
Table S1. Plasmids and strains used in this study Genotype or phenotype
Induction
Source or reference
Plasmids pKG116
Expression vector, Camr
-
1
pHP2
Tar expression plasmid with added NotI and deleted
-
2
2 µM sodium
3
NdeI restriction site, Ampr pPA791
Tar∆(44-183)/T303I expression plasmid, Camr
salicylate pVS1092
Tar expression plasmid, Camr
2 µM sodium
4
salicylate pVS88
CheY-EYFP / CheZ-ECFP expression plasmid, Ampr
50 µM IPTG
5
pOB30
GFP expression plasmid, Ampr
50 µM IPTG
Gift of Olga Besharova
pSB1
pSB2
pSB3
pSB4
pSB5
pSB6
Hybrid Tar[1-43]-McpC[49-270]-Tar[184-553]
2 µM sodium
expression plasmid, pKG116 derivative
salicylate
Hybrid McpC[1-286]-Tar[200-553] expression plasmid,
2 µM sodium
pKG116 derivative
salicylate
Hybrid McpC[1-350]-Tar[267-553] expression plasmid,
2 µM sodium
pKG116 derivative
salicylate
Hybrid McpC[1-340] and Tar[257-553] expression
2 µM sodium
plasmid, pKG116 derivative
salicylate
Hybrid McpC[1-472]-Tar[361-417]-McpC[530-655]
2 µM sodium
expression plasmid, pKG116 derivative
salicylate
Hybrid McpC[1-350]-YSYEH-Tar[272-553] expression
2 µM sodium
plasmid, pKG116 derivative
salicylate 16
This study
This study
This study
This study
This study
This study
pSB7
pSB8
pSB9
pSB10
pSB11
pSB12
Hybrid Tar[1-43]-CitA[55-173]-Tar[184-553]
2 µM sodium
expression plasmid, pKG116 derivative
salicylate
Hybrid CitA[1-193]-Tar[204-553] expression plasmid,
2 µM sodium
pKG116 derivative
salicylate
Hybrid ECA0434[1-385]-Tar[267-553] expression
2 µM sodium
plasmid, pKG116 derivative
salicylate
Hybrid McpS[1-295]-Tar[200-553] expression plasmid,
0.7 µM sodium
pKG116 derivative
salicylate
Hybrid NarQ[1-217]-Tar[257-553] expression plasmid,
2 µM sodium
pKG116 derivative
salicylate
Hybrid McpG[1-348]-Tar[268-553] expression plasmid,
0.8 µM sodium
pKG116 derivative
salicylate
This study
This study
This study
This study
This study
6
Strains UU1250
E. coli RP437 ΔaerΔtsrΔ(tar-tap)Δtrg
-
7
VS181
E. coli RP437 Δ(cheYcheZ)ΔaerΔtsrΔ(tar-tap)Δtrg
-
5
VH1
E. coli RP437
-
8
Δ(cheRcheB)Δ(cheYcheZ)ΔaerΔtsrΔ(tar-tap)Δtrg
17
Table S2. List of all compounds screened for McpC350Tar267
Name
Response of hybrid receptor
Putrescine
Nb
N-phthaloyl-Lglutamic acid N-acetyl-L-glutamic acid Glycyl-L-aspartic acid D,L-lactamide Glycyl-Lmethionine
Response of Tara
Name D-2-phospho-glyceric acid D-3-phospho-glyceric acid
N N
O-phospho-D-serine
N N N
O-phospho-Lthreonine O-phospho-L-tyrosine O-phospho-Dtyrosine O-phospho-Dthreonine O-phospho-L-serine m-Hydroxy-phenyl acetic acid L-methionine sulfoxide Hydroxylamine Methylamine Parabanic acid
Response of hybrid receptor N N N N N N
Adenine
N
L-ornithine
N
L-cysteinyl-glycine
N
Ala-Asp
N
L-homoserine L-citrulline Ala-Gly
-c N N
-Hydroxy-butyric acid
N
n-Butylamine
N
-Hydroxy-butyric acid
N
Ethylamine
N
N N
Ethanolamine Ethylenediamine
N N
N
Glucuronamide
N
N
Agmatine
N
Histamine
N
-Phenylethyl-amine
N
Tyramine
N
-Keto-valeric acid Ala-Gln 3-Hydroxy-2butanone -Amino-n-caproica cid -Amino-n-valeric acid p-Hydroxy-phenyl acetic acid D,L--amino-nbutyric acid
-
N
N
N -
N 18
N N N N N N N
Response of Tara
-Amino-n-butyric acid
N
Acetamide
N
-Amino-n-valeric acid
N
Formamide
N
D-alanine
-
D-aspartic acid D-methionine D-asparagine D-threonine D-cysteine D-glutamic acid D-lysine D-valine D-serine Inosine Xanthine Uridine Acetamide D,L-carnitine D,L-octopamine 2,3-Butanediol N-acetyl-L-cysteine
N N N N N N N N N N N N N N N N N
-Hydroxy-glutaric acid--lactone
N
a
D,L--amino-caprylic acid Adenosine Cytidine Cytosine Guanine Guanosine Thymine Thymidine Uracil Allantoin Alloxan Uric acid Xanthosine Ala-Glu Sec-butylamine Dihydroxyaceton 2,3-Butanone S-methyl-L-cysteine
N
-Hydroxy-butyric acid
N N N N N N N N N N N N N N N N N N N
Measured only for compounds that induce a chemotactic response via the hybrid receptor. bNo
chemotactic response. cRepellent response.
19
Table S3. List of all compounds screened for McpG348Tar268
Name
Response of hybrid receptor
L-aspartic acid
Nb
L-glutamic acid
N
L-alanine L-asparagine L-cysteine L-arginine L-glutamine Glycine L-histidine L-isoleucine L-leucine L-lysine L-methionine
+c N + N N N N N N N N
L-phenylalanine
N
L-proline L-serine L-threonine L-valine L-tryptophan L-tyrosine L-homoserine L-ornithine D-alanine D-aspartic acid D-methionine D-asparagine D-threonine D-cysteine D-glutamic acid D-lysine D-valine D-serine N-acetyl-Lglutamic acid L-citrulline
+ N N N N N + N N N N N N N N N N N
Response of Tara
+ +
Name Urea N-phthaloyl-Lglutamic acid L-pyroglutamic acid Hydroxylamine Methylamine N-amylamine N-butylamine Ethylamine Ethanolamine Ethylenediamine Putrescine Agmatine Histamine -Phenylethylamine Tyramine Acetamide Formamide Adenine Adenosine Cytidine Cytosine Guanine Guanosine Thymine Thymidine Uracil Uridine Inosine Xanthine Xanthosine Uric acid Alloxan
N
N
Response of hybrid receptor N N N N N N N N N + + + N N N N N N N N N N N N N N N N N N N N
N
Allantoin
N
N
Parabanic acid
N
20
Response of Tara
N N N
D,L--amino-nbutyric acid D,L--aminocaprylic acid Glucuronamide a
-Amino-n-caproic acid
N
-Amino-n-valeric acid D,L-lactamide
N N
N N N
Measured only for compounds that induce a chemotactic response via the hybrid receptor. bNo
chemotactic response. cAttractant response.
21
Table S4. List of all compounds screened for McpS295Tar200
Name
Response of hybrid receptor
Name
Response of hybrid receptor
L-aspartic acid L-glutamic acid
+b Nc
D-asparagine D-glutamic acid
N N
L-alanine
N
D,L--amino-nbutyric acid
N
L-asparagine
N
-Amino-n-butyric acid
N
L-cysteine
N
-Amino-n-caproica cid
N
L-arginine
N
-Amino-valeric acid
N
L-glutamine
N
Glycine L-histidine
N N
L-isoleucine
N
-Hydroxy-butyric acid
N
L-leucine
N
-Hydroxy-butyric acid
N
L-lysine
N
L-methionine
N
L-phenylalanine L-proline L-serine L-threonine
N N N N
L-valine
N
L-tryptophan
N
L-tyrosine
N
Methyl-pyruvate
N
Oxaloacetic acid
N
D,L--hydroxybutyric acid D-aspartic acid Itaconic acid
Response of Tara +
-Amino-n-valeric acid Propionic acid Caproic acid
-Keto-valeric acid -Hydroxy-butyric acid Citraconic acid L-tartaric acid Quinolinic acid Nicotinic acid -Amino-levulinic acid D,L-mevalonic acid
N N +
N N N N N N N
N
N
Methyl-pyruvate
N
N +
Oxaloacetic acid Valeric acid
N +
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N
N
-Hydroxy-glutaric acid--lactone Acetoacetic acid Mono methylsuccinate
N
Response of Tara
+
N
N
N
-Keto-butyric acid
N
D-tartaric acid
+
N
a
Measured only for compounds that induce chemotactic response via the hybrid receptor.
b
Attractant response. cNo chemotactic response.
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Supplementary References 1. Buron-Barral, M. C., Gosink, K. K., and Parkinson, J. S. (2006) Loss- and gain-of-function mutations in the F1-HAMP region of the Escherichia coli aerotaxis transducer Aer. J. Bacteriol. 188, 3477-3486. 2. Pham, H. T., and Parkinson, J. S. (2011) Phenol sensing by Escherichia coli chemoreceptors: a nonclassical mechanism. J. Bacteriol. 193, 6597-6604. 3. Gosink, K. K., Buron-Barral, M. C., and Parkinson, J. S. (2006) Signaling interactions between the aerotaxis transducer Aer and heterologous chemoreceptors in Escherichia coli. J. Bacteriol. 188, 3487-3493. 4. Yang, Y., and Sourjik, V. (2012) Opposite responses by different chemoreceptors set a tunable preference point in Escherichia coli pH taxis. Mol. Microbiol. 86, 1482-1489. 5. Sourjik, V., and Berg, H. C. (2004) Functional interactions between receptors in bacterial chemotaxis. Nature 428, 437-441. 6. Reyes-Darias, J. A., Garcia, V., Rico-Jimenez, M., Corral-Lugo, A., Lesouhaitier, O., Juarez-Hernandez, D., Yang, Y., Bi, S., Feuilloley, M., Munoz-Rojas, J., Sourjik, V., and Krell, T. (2015) Specific gamma-aminobutyrate chemotaxis in pseudomonads with different lifestyle. Mol. Microbiol. 97, 488-501. 7. Ames, P., Studdert, C. A., Reiser, R. H., and Parkinson, J. S. (2002) Collaborative signaling by mixed chemoreceptor teams in Escherichia coli. Proc. Natl. Acad. Sci. U.S.A. 99, 7060-7065. 8. Oleksiuk, O., Jakovljevic, V., Vladimirov, N., Carvalho, R., Paster, E., Ryu, W. S., Meir, Y., Wingreen, N. S., Kollmann, M., and Sourjik, V. (2011) Thermal robustness of signaling in bacterial chemotaxis, Cell 145, 312-321.
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