Record ID
xffwr-bf334
Publication date
Jan 5, 2026
Depositors
Gustavo Fuertes
,
Phuong Ngoc Pham
,
Giusy Finocchiaro
,
Jiří Homola
Associated publication
Article, Front-illuminated surface plasmon resonance biosensor for the study of light-responsive proteins and their interactions
License
Funding references
Time-resolved vibrational spectroscopy of proteins assisted by genetically encoded non-canonical amino acids (Czech Science Foundation (GACR): 24-11819S),
Innovation of Czech Infrastructure for Integrative Structural Biology (European Regional Development Fund: CZ.02.01.01/00/23_015/0008175),
CIISB, Instruct-CZ Centre of Instruct-ERIC EU consortium (MEYS CR: LM2023042)
Method
Bio-layer interferometry (BLI)
Entities of interest
Entity of interest 1
Name
EL222
Type
Polymer
Molecular weight
23157.0 g/mol
Source organism
Erythrobacter litoralis
Sequence
GADDTRVEVQ PPAQWVLDL I EASPIASV VS DPRLADN PLI AINQAF
TDLT GYSEE ECVGR NCRF LAGSGT EPW LTDKIRQ GV REHKPVLV E
ILNYKKDGT PFRNAVLVAP IYDDDDELL Y FLGSQVEV DD DQPNMGM
ARR ERAAEM LKTL SPRQL EVTTL VASG LRNKEV AAR LGLSEKT VK
MHRGLVME K LNLKTSADL VRIAVEAGIA
TDLT GYSEE ECVGR NCRF LAGSGT EPW LTDKIRQ GV REHKPVLV E
ILNYKKDGT PFRNAVLVAP IYDDDDELL Y FLGSQVEV DD DQPNMGM
ARR ERAAEM LKTL SPRQL EVTTL VASG LRNKEV AAR LGLSEKT VK
MHRGLVME K LNLKTSADL VRIAVEAGIA
Entity of interest 2
Name
dsDNA20
Type
Polymer
Molecular weight
16625.0 g/mol
Sequence
Forward: 5 '- TTT TTT TTT TTT T TA GGT AGC CTT TAG T
CC ATG -3' Reverse: 5’- CAT GG A CTA AAG GCT ACC TA
-3’
CC ATG -3' Reverse: 5’- CAT GG A CTA AAG GCT ACC TA
-3’
Modifications
Chemical
Type
Biotin
Position
5' (forward strand)
Entity of interest 3
Name
Consensus b-ssDNA
Type
Polymer
Molecular weight
10515.0 g/mol
Sequence
5'- TTT TT T TTT TTT TTA GGT AG C CTT TAG TCC ATG -3
'
'
Modifications
Chemical
Type
Biotin
Position
5'
Entity of interest 4
Name
Complementary consensus ssDNA
Type
Polymer
Molecular weight
6111.0 g/mol
Sequence
5’- CAT GG A CTA AAG GCT ACC TA -3
Results
Result 1
Name
ka,D-DNA (association rate of EL222 dimer interaction with DNA)
Value
2000.0 M^-1 s^-1
Result type
Association rate kOn
Entity 1
Name
EL222
copy number
2.0
Entity 2
Name
dsDNA20
copy number
1.0
Result 2
Name
kd,D-DNA (dissociation constant of EL222 dimer interaction with DNA)
Value
0.002 s^-1
Result type
Dissociation rate kOff
Entity 1
Name
EL222
copy number
2.0
Entity 2
Name
dsDNA20
copy number
1.0
Result 3
Name
KD,D-DNA (equilibrium dissociation constant of EL222 dimer interaction with DNA)
Value
1.0 µM
Result type
Constant of dissociation KD
Entity 1
Name
EL222
copy number
2.0
Entity 2
Name
dsDNA20
copy number
1.0
Chemical environments (buffers)
Name
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Constituents
2-(N-morpholino)ethanesulfonic acid
50.0
mM
Sodium chloride
100.0
mM
Bovine Serum Albumin (BSA)
250.0
µg/mL
pH
7.4
Name
PBS
Constituents
KH2PO4
1.4
mM
Na2HPO4
8.0
mM
KCl
2.7
mM
NaCl
137.0
mM
pH
7.4
Measurements
Experimental setup
Experiment type
Affinity
Measurement Protocol
First baseline (before illumination)
Baseline
880.0 seconds
Immobilization of biotinylated forward-ssDNA (before illumination)
Load
90.0 seconds
Second baseline (before illumination)
Baseline
180.0 seconds
Immobilization of reverse-ssDNA for in situ annealing of dsDNA (before illumination)
Load
90.0 seconds
Third baseline (immediately after illumination)
Baseline
180.0 seconds
EL222-DNA association (3 minutes after illumination)
Association
301.0 seconds
EL222-DNA dissociation (8 minutes after illumination)
Dissociation
1600.0 seconds
Measurement 1
Name
Set 2: EL222 20 uM third baseline
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 2
Name
Set 2: EL222 16 uM third baseline
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 3
Name
Set 2: EL222 12 uM third baseline
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 4
Name
Set 2: EL222 8 uM third baseline
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 5
Name
Set 2: EL222 6 uM third baseline
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 6
Name
Set 2: EL222 4 uM third baseline
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 7
Name
Set 2: EL222 2 uM third baseline
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 8
Name
Set 2: EL222 0 uM association (third baseline)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 9
Name
Set 2: EL222 20 uM association (3 minutes after illumination)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 10
Name
Set 2: EL222 16 uM association (3 minutes after illumination)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 11
Name
Set 2: EL222 12 uM association (3 minutes after illumination)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 12
Name
Set 2: EL222 8 uM association (3 minutes after illumination)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 13
Name
Set 2: EL222 6 uM association (3 minutes after illumination)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 14
Name
Set 2: EL222 4 uM association (3 minutes after illumination)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 15
Name
Set 2: EL222 2 uM association (3 minutes after illumination)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 16
Name
Set 2: EL222 0 uM association (3 minutes after illumination)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 17
Name
Set 2: EL222 20 uM dissociation
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 18
Name
Set 2: EL222 16 uM dissociation
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 19
Name
Set 2: EL222 12 uM dissociation
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 20
Name
Set 2: EL222 8 uM dissociation
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 21
Name
Set 2: EL222 6 uM dissociation
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 22
Name
Set 2: EL222 4 uM dissociation
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 23
Name
Set 2: EL222 2 uM dissociation
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 24
Name
Set 2: EL222 0 uM dissociation
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 25
Name
Set 1: First baseline
Chemical environment
PBS
Measurement 26
Name
Set 1: First baseline
Chemical environment
PBS
Measurement 27
Name
Set 1: First baseline
Chemical environment
PBS
Measurement 28
Name
Set 1: First baseline
Chemical environment
PBS
Measurement 29
Name
Set 1: First baseline
Chemical environment
PBS
Measurement 30
Name
Set 1: First baseline
Chemical environment
PBS
Measurement 31
Name
Set 1: Immobilization of biotinylated DNA (forward strand)
Chemical environment
PBS
Measurement 32
Name
Set 1: Immobilization of biotinylated DNA (forward strand)
Chemical environment
PBS
Measurement 33
Name
Set 1: Immobilization of biotinylated DNA (forward strand)
Chemical environment
PBS
Measurement 34
Name
Set 1: Immobilization of biotinylated DNA (forward strand)
Chemical environment
PBS
Measurement 35
Name
Set 1: Immobilization of biotinylated DNA (forward strand)
Chemical environment
PBS
Measurement 36
Name
Set 1: Immobilization of biotinylated DNA (forward strand)
Chemical environment
PBS
Measurement 37
Name
Set 1: Second baseline
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 38
Name
Set 1: Second baseline
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 39
Name
Set 1: Second baseline
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 40
Name
Set 1: Second baseline
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 41
Name
Set 1: Second baseline
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 42
Name
Set 1: Second baseline
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 43
Name
Set 1: Immobilization of DNA (reverse strand)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 44
Name
Set 1: Immobilization of DNA (reverse strand)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 45
Name
Set 1: Immobilization of DNA (reverse strand)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 46
Name
Set 1: Immobilization of DNA (reverse strand)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 47
Name
Set 1: Immobilization of DNA (reverse strand)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Measurement 48
Name
Set 1: Immobilization of DNA (reverse strand)
Chemical environment
MES 50 mM, NaCl 150 mM, BSA 250 µg/mL
Instrument
Manufacturer
Sartorius
Name
Octet R8 (Eight-Channel System)
Files (6)
Derived measurement data
Set 2 (after illumination): Baseline-EL222/DNA association-EL222/DNA dissociation (EL222 concentrations 20-16-8-4 uM). BLI sensor response (nm) vs. time (s)
Download BLI_DNA_EL222_3_MINUTES_AFTER_ILLUMINATION_DATA.csv
Raw measurement data
Set 2 (after illumination): Baseline-EL222/DNA association-EL222/DNA dissociation (EL222 concentrations 12-6 uM)
Download Preprocessed Report_2025_07_15 11_36_39_DNA_EL222_12-6-uM.xlsx
Raw measurement data
Zip folder containing the original raw data files (frd format)
Download 2025_07_14.zip
Derived measurement data
Set 2 (after illumination): Baseline-EL222/DNA association-EL222/DNA dissociation (EL222 concentrations 20-16-8-4 uM). Fitted BLI sensor response (nm) vs. time (s)
Download BLI_DNA_EL222_3_MINUTES_AFTER_ILLUMINATION_FIT.csv
Raw measurement data
Set 2 (after illumination): Baseline-EL222/DNA association-EL222/DNA dissociation (EL222 concentrations 20-16-8-4-2-0 uM)
Download Preprocessed Report_2025_07_15 11_00_18_DNA_EL222_20-16-8-4-2-uM.xlsx
Raw measurement data
Set 1 (before illumination): Baseline-Loading (biotinylated forward ss DNA)-Baseline-Loading (reverse ssDNA)
Download Preprocessed Report_2025_07_15 11_51_50_DNA_500_nM_fw_100_nM_rv.xlsx