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- PDB-6am7: Engineered tryptophan synthase b-subunit from Pyrococcus furiosus... -

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Basic information

Entry
Database: PDB / ID: 6am7
TitleEngineered tryptophan synthase b-subunit from Pyrococcus furiosus, PfTrpB2B9
ComponentsTryptophan synthase beta chain 1
KeywordsBIOSYNTHETIC PROTEIN / PLP Type II / Tryptophan Synthase / Engineered / Allostery
Function / homology
Function and homology information


tryptophan synthase activity / tryptophan synthase
Similarity search - Function
Tryptophan synthase, beta chain, conserved site / Tryptophan synthase, beta chain / Tryptophan synthase beta chain/beta chain-like / Tryptophan synthase beta chain pyridoxal-phosphate attachment site. / Rossmann fold - #1100 / Pyridoxal-phosphate dependent enzyme / Tryptophan synthase beta subunit-like PLP-dependent enzyme / Pyridoxal-phosphate dependent enzyme / Rossmann fold / 3-Layer(aba) Sandwich / Alpha Beta
Similarity search - Domain/homology
PHOSPHATE ION / Tryptophan synthase beta chain 1
Similarity search - Component
Biological speciesPyrococcus furiosus (archaea)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.47 Å
AuthorsBuller, A.R. / van Roye, P.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)F32GM110851 United States
CitationJournal: J. Am. Chem. Soc. / Year: 2018
Title: Directed Evolution Mimics Allosteric Activation by Stepwise Tuning of the Conformational Ensemble.
Authors: Buller, A.R. / van Roye, P. / Cahn, J.K.B. / Scheele, R.A. / Herger, M. / Arnold, F.H.
History
DepositionAug 9, 2017Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 16, 2018Provider: repository / Type: Initial release
Revision 1.1Jun 20, 2018Group: Data collection / Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.title
Revision 1.2Mar 23, 2022Group: Author supporting evidence / Database references / Derived calculations
Category: database_2 / pdbx_audit_support ...database_2 / pdbx_audit_support / pdbx_struct_conn_angle / struct_conn / struct_conn_type
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_audit_support.funding_organization / _pdbx_audit_support.grant_number / _pdbx_struct_conn_angle.ptnr1_auth_asym_id / _pdbx_struct_conn_angle.ptnr1_auth_comp_id / _pdbx_struct_conn_angle.ptnr1_auth_seq_id / _pdbx_struct_conn_angle.ptnr1_label_alt_id / _pdbx_struct_conn_angle.ptnr1_label_asym_id / _pdbx_struct_conn_angle.ptnr1_label_atom_id / _pdbx_struct_conn_angle.ptnr1_label_comp_id / _pdbx_struct_conn_angle.ptnr1_label_seq_id / _pdbx_struct_conn_angle.ptnr1_symmetry / _pdbx_struct_conn_angle.ptnr2_auth_asym_id / _pdbx_struct_conn_angle.ptnr2_auth_seq_id / _pdbx_struct_conn_angle.ptnr2_label_asym_id / _pdbx_struct_conn_angle.ptnr2_symmetry / _pdbx_struct_conn_angle.ptnr3_auth_asym_id / _pdbx_struct_conn_angle.ptnr3_auth_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_seq_id / _pdbx_struct_conn_angle.ptnr3_label_alt_id / _pdbx_struct_conn_angle.ptnr3_label_asym_id / _pdbx_struct_conn_angle.ptnr3_label_atom_id / _pdbx_struct_conn_angle.ptnr3_label_comp_id / _pdbx_struct_conn_angle.ptnr3_label_seq_id / _pdbx_struct_conn_angle.ptnr3_symmetry / _pdbx_struct_conn_angle.value / _struct_conn.conn_type_id / _struct_conn.id / _struct_conn.pdbx_dist_value / _struct_conn.pdbx_leaving_atom_flag / _struct_conn.pdbx_ptnr1_label_alt_id / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr1_symmetry / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id / _struct_conn.ptnr2_label_seq_id / _struct_conn.ptnr2_symmetry / _struct_conn_type.id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Tryptophan synthase beta chain 1
B: Tryptophan synthase beta chain 1
C: Tryptophan synthase beta chain 1
D: Tryptophan synthase beta chain 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)175,01615
Polymers174,4754
Non-polymers54111
Water15,385854
1
A: Tryptophan synthase beta chain 1
B: Tryptophan synthase beta chain 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)87,4967
Polymers87,2372
Non-polymers2595
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area4320 Å2
ΔGint-68 kcal/mol
Surface area26030 Å2
MethodPISA
2
C: Tryptophan synthase beta chain 1
D: Tryptophan synthase beta chain 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)87,5198
Polymers87,2372
Non-polymers2826
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area4450 Å2
ΔGint-75 kcal/mol
Surface area26420 Å2
MethodPISA
Unit cell
Length a, b, c (Å)83.952, 107.822, 159.772
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number19
Space group name H-MP212121

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Components

#1: Protein
Tryptophan synthase beta chain 1


Mass: 43618.719 Da / Num. of mol.: 4
Mutation: I16V, E17G, I68V, F95L, F274S, T292S, T321A, V384A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus furiosus (strain ATCC 43587 / DSM 3638 / JCM 8422 / Vc1) (archaea)
Strain: ATCC 43587 / DSM 3638 / JCM 8422 / Vc1 / Gene: trpB1, PF1706 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q8U093, tryptophan synthase
#2: Chemical
ChemComp-NA / SODIUM ION


Mass: 22.990 Da / Num. of mol.: 7 / Source method: obtained synthetically / Formula: Na
#3: Chemical
ChemComp-PO4 / PHOSPHATE ION / Phosphate


Mass: 94.971 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: PO4
#4: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 854 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.07 Å3/Da / Density % sol: 40.64 %
Crystal growTemperature: 298 K / Method: vapor diffusion, sitting drop / pH: 7.85 / Details: 15-25% PEG3350 and 0.1 M Na HEPES pH 7.85

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: SSRL / Beamline: BL12-2 / Wavelength: 0.9795 Å
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Jul 9, 2015
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9795 Å / Relative weight: 1
ReflectionResolution: 1.47→40 Å / Num. obs: 243063 / % possible obs: 98.4 % / Redundancy: 4.3 % / CC1/2: 0.999 / Rpim(I) all: 0.031 / Net I/σ(I): 12.1
Reflection shellResolution: 1.47→1.49 Å / Redundancy: 4.8 % / Num. unique obs: 9310 / CC1/2: 0.284 / % possible all: 77.3

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Processing

Software
NameVersionClassification
REFMAC5.8.0158refinement
XDSdata reduction
Aimlessdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5DVZ
Resolution: 1.47→40 Å / Cor.coef. Fo:Fc: 0.964 / Cor.coef. Fo:Fc free: 0.958 / SU B: 4.003 / SU ML: 0.07 / Cross valid method: THROUGHOUT / ESU R: 0.079 / ESU R Free: 0.075 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflectionSelection details
Rfree0.21368 12028 5 %RANDOM
Rwork0.19926 ---
obs0.19998 230004 98.35 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK
Displacement parametersBiso mean: 24.493 Å2
Baniso -1Baniso -2Baniso -3
1-1.92 Å20 Å20 Å2
2---0.38 Å20 Å2
3----1.53 Å2
Refinement stepCycle: 1 / Resolution: 1.47→40 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms11567 0 27 854 12448
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0060.01911917
X-RAY DIFFRACTIONr_bond_other_d0.0020.0211195
X-RAY DIFFRACTIONr_angle_refined_deg1.1411.97516162
X-RAY DIFFRACTIONr_angle_other_deg0.865325874
X-RAY DIFFRACTIONr_dihedral_angle_1_deg5.40551554
X-RAY DIFFRACTIONr_dihedral_angle_2_deg36.69823.875480
X-RAY DIFFRACTIONr_dihedral_angle_3_deg11.523151947
X-RAY DIFFRACTIONr_dihedral_angle_4_deg14.9041563
X-RAY DIFFRACTIONr_chiral_restr0.0590.21783
X-RAY DIFFRACTIONr_gen_planes_refined0.0040.02113439
X-RAY DIFFRACTIONr_gen_planes_other0.0010.022392
X-RAY DIFFRACTIONr_nbd_refined
X-RAY DIFFRACTIONr_nbd_other
X-RAY DIFFRACTIONr_nbtor_refined
X-RAY DIFFRACTIONr_nbtor_other
X-RAY DIFFRACTIONr_xyhbond_nbd_refined
X-RAY DIFFRACTIONr_xyhbond_nbd_other
X-RAY DIFFRACTIONr_metal_ion_refined
X-RAY DIFFRACTIONr_metal_ion_other
X-RAY DIFFRACTIONr_symmetry_vdw_refined
X-RAY DIFFRACTIONr_symmetry_vdw_other
X-RAY DIFFRACTIONr_symmetry_hbond_refined
X-RAY DIFFRACTIONr_symmetry_hbond_other
X-RAY DIFFRACTIONr_symmetry_metal_ion_refined
X-RAY DIFFRACTIONr_symmetry_metal_ion_other
X-RAY DIFFRACTIONr_mcbond_it0.1841.0746144
X-RAY DIFFRACTIONr_mcbond_other0.1841.0746143
X-RAY DIFFRACTIONr_mcangle_it0.3451.6097680
X-RAY DIFFRACTIONr_mcangle_other0.3451.6097681
X-RAY DIFFRACTIONr_scbond_it0.1171.0835773
X-RAY DIFFRACTIONr_scbond_other0.1151.0785758
X-RAY DIFFRACTIONr_scangle_it
X-RAY DIFFRACTIONr_scangle_other0.2181.6118445
X-RAY DIFFRACTIONr_long_range_B_refined4.11713.45513474
X-RAY DIFFRACTIONr_long_range_B_other4.06212.95913281
X-RAY DIFFRACTIONr_rigid_bond_restr
X-RAY DIFFRACTIONr_sphericity_free
X-RAY DIFFRACTIONr_sphericity_bonded
LS refinement shellResolution: 1.469→1.507 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0.372 654 -
Rwork0.362 13674 -
obs--79.53 %
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL112)L122)L132)L222)L232)L332)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T112)T122)T132)T222)T232)T332)Origin x (Å)Origin y (Å)Origin z (Å)
10.1178-0.077-0.68712.3790.68094.80950.0695-0.00960.0269-0.422-0.0019-0.0556-0.69910.4646-0.06760.2373-0.1320.0290.21580.04950.042710.348649.7771-1.5442
20.8891-0.03510.03722.27270.54441.47380.02470.03850.01560.01770.0076-0.1676-0.18370.1712-0.03230.0565-0.01580.00650.1508-0.01310.126112.113336.498223.8584
30.8446-0.5569-0.35211.76520.81622.05470.07810.06380.094-0.1774-0.09130.0121-0.4501-0.02570.01320.2215-0.00170.01810.11120.00790.11623.829150.2715.5302
48.25894.36124.25734.96493.09565.65240.04590.3860.19350.0686-0.14690.1951-0.4355-0.19920.1010.23250.08530.03480.11-0.0050.0365-5.896954.088313.4084
52.6993-0.2871-1.15051.81790.98554.7625-0.11550.4443-0.1926-0.2462-0.1720.2368-0.5148-0.97910.28750.22190.1134-0.02050.2274-0.06250.0441-8.03151.697210.7846
60.48240.15390.14041.1173-0.26891.7496-0.01550.1052-0.0772-0.18440.0234-0.1701-0.03530.1608-0.00790.0789-0.01240.03380.14-0.01880.119612.106133.688411.7126
73.72584.229-2.38675.5471-3.96654.0797-0.17980.1728-0.0064-0.50090.2690.12330.4016-0.3615-0.08920.23680.0012-0.10760.1555-0.02270.0649-4.374131.17810.0862
80.1562-0.2061.01080.3339-1.56157.45850.0156-0.0795-0.05390.01770.25060.0649-0.1644-0.991-0.26620.17250.0167-0.07380.35530.0340.073-6.704233.54262.3569
90.84090.169-0.17080.9012-0.52661.6166-0.00020.0661-0.1212-0.1520.0122-0.01010.1211-0.0318-0.0120.062-0.0141-0.01520.1128-0.02490.11682.908324.68116.4099
104.1161-2.9415-0.60196.43061.19297.26430.0530.1343-0.0147-0.253-0.10230.5366-0.0584-0.19640.04920.1708-0.02080.00780.24110.00830.213-14.709130.920624.0771
110.8666-0.8858-0.52882.26850.79183.8285-0.0608-0.1392-0.12090.22590.117-0.08620.36360.1836-0.05630.10590.0263-0.02730.13970.00630.07225.923128.945461.1954
120.52440.2919-0.21520.75750.0871.21850.0104-0.0259-0.029-0.00330.019-0.02560.03180.1098-0.02940.05560.0124-0.01730.1271-0.01120.13965.142635.177440.0758
132.933-0.5973-1.36361.7550.52774.77910.07310.03810.2356-0.1522-0.05770.1873-0.1939-0.47-0.01540.0345-0.0125-0.0260.1259-0.01670.1256-13.062129.908738.9553
147.5325-5.81982.28137.6471-0.11723.4077-0.1053-0.69660.0280.494-0.04680.13120.2446-0.82590.15210.126-0.0791-0.00150.2248-0.04070.018-10.259325.917954.1817
150.44540.2106-0.20211.3428-0.5481.4228-0.01-0.10440.00270.05170.0145-0.1040.06150.1861-0.00450.04730.0032-0.02050.1406-0.01480.12068.111136.592349.5637
162.1206-0.1456-0.05641.1645-0.20851.4448-0.0042-0.14380.16430.05670.0393-0.1031-0.29180.1325-0.03510.1249-0.0203-0.01620.1351-0.0360.13537.0451.365148.9914
172.8187-1.62971.53912.6883-2.83.7622-0.2105-0.14110.20240.35610.26320.0193-0.3808-0.3916-0.05270.07180.06870.0110.17440.00320.0514-6.475944.904559.8731
181.2757-1.33442.87732.0555-2.44716.97050.0106-0.7314-0.36680.14320.72860.49360.1561-1.6742-0.73920.06190.02030.00360.47260.16620.1638-10.798344.614152.4923
190.6115-0.08910.17441.0163-0.43011.8437-0.0022-0.06310.1590.01190.0049-0.0606-0.37870.0362-0.00270.12020.00580.00810.0954-0.02610.12831.549954.184743.7457
2010.629-4.3071.69866.0566-0.62255.79020.13470.1529-0.1487-0.4130.1150.3931-0.1417-0.4392-0.24970.13340.0214-0.05120.1613-0.00350.1319-11.833946.296532.6487
216.28040.59870.338410.27930.3584.4519-0.09460.4268-0.0526-0.45190.29120.2535-0.086-0.2908-0.19660.137-0.0317-0.02240.14850.04280.073435.859719.263314.5012
220.75330.38810.54211.01080.73092.07890.0195-0.0316-0.0448-0.0156-0.0340.07850.0672-0.22160.01450.0188-0.0025-0.01250.10790.0150.161127.272213.33932.8966
230.71920.58180.32871.80760.42821.01220.0286-0.0102-0.02230.06470.0253-0.04990.02840.0378-0.05390.0569-0.0007-0.00920.14110.0140.155938.933217.038739.2747
242.7766-0.1755-0.91021.63590.10182.33130.02540.01220.0676-0.00540.0001-0.17640.27380.2443-0.02550.08670.0473-0.03810.07340.01460.127344.31760.850839.2204
256.53830.25420.75421.5311-0.3180.9188-0.15880.5219-0.1598-0.37110.0789-0.15320.3620.35690.080.26210.06930.01220.19790.01660.098242.9664-1.152324.7601
260.72780.410.44171.18010.58341.4272-0.05050.03990.0964-0.05160.04160.0613-0.1850.04080.0090.0646-0.0192-0.00550.11750.01230.154336.469324.471531.3301
277.16451.77751.5172.10260.89332.8494-0.06070.22280.1394-0.07370.113-0.2608-0.11780.4632-0.05220.0762-0.03080.02640.1732-0.03130.090950.871617.723219.4872
280.8251-1.6249-0.99566.74687.901413.5080.33750.4194-0.0768-0.49390.0928-0.24380.79840.9345-0.43030.5050.17750.00370.4555-0.14730.059454.233113.095925.346
291.61690.0211.5431.0680.09952.9239-0.07310.15410.1023-0.18520.0975-0.1533-0.21830.3656-0.02440.0518-0.06150.03570.1415-0.01340.115949.08426.343729.7733
300.96040.2254-0.02571.97980.36281.476-0.0415-0.00120.02750.01980.0437-0.1684-0.02450.2413-0.00220.0423-0.0421-0.00460.1605-0.01710.145449.648327.776941.1902
310.21050.40980.48992.5945-0.94384.2147-0.0285-0.0460.07760.36710.01940.0441-0.63980.06380.0090.1775-0.00490.00870.1485-0.07790.120543.836328.641681.5339
321.17670.51190.36341.50890.25261.81090.0041-0.05650.05820.02640.02420.1347-0.0603-0.1301-0.02830.08670.00750.01450.15170.01060.130734.802519.954958.3008
331.5368-0.25320.14223.5931.41843.4884-0.1007-0.02520.00580.34730.10140.0018-0.05180.2921-0.00070.1125-0.06240.0180.1336-0.03260.07951.156827.278364.038
341.4005-0.18790.96775.7914-0.28266.386-0.08050.0687-0.16830.44690.3882-0.37180.44081.2091-0.30770.10530.0873-0.02430.3531-0.10310.062759.39720.321960.9947
352.5965-2.8822-0.607112.78093.87881.22650.4734-0.1305-0.27650.272-0.1723-0.80880.1844-0.008-0.30110.44950.0459-0.08360.39890.07780.171960.631520.908274.8553
360.99960.56320.53691.32440.58361.9606-0.0478-0.08320.12320.09190.05710.0293-0.23420.0113-0.00930.11880.0070.01090.1526-0.00350.098140.216724.539770.1236
371.5821-0.29660.07491.88360.1311.94270.0211-0.1451-0.01540.17040.03990.19090.1966-0.3061-0.0610.111-0.04330.00960.20060.04020.102529.625311.176668.808
380.6908-0.4357-0.51011.84493.30616.57280.0432-0.1945-0.11310.2830.1751-0.10550.58030.5361-0.21830.18520.053-0.06130.23360.03810.044348.12556.138277.2191
390.70740.05560.28590.6080.88822.93160.0564-0.1219-0.17590.1909-0.00710.07850.5945-0.1137-0.04930.1817-0.0179-0.01960.10.05410.103936.21333.541565.9632
404.3418-0.2636-1.6251.66850.36242.93140.01820.0038-0.27880.01680.0444-0.06760.40650.0958-0.06270.1350.0234-0.04730.10.02140.070639.02275.498257.1436
Refinement TLS group
IDRefine-IDRefine TLS-IDAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1A1 - 29
2X-RAY DIFFRACTION2A30 - 79
3X-RAY DIFFRACTION3A80 - 121
4X-RAY DIFFRACTION4A122 - 132
5X-RAY DIFFRACTION5A133 - 180
6X-RAY DIFFRACTION6A181 - 254
7X-RAY DIFFRACTION7A255 - 278
8X-RAY DIFFRACTION8A279 - 305
9X-RAY DIFFRACTION9A306 - 378
10X-RAY DIFFRACTION10A379 - 384
11X-RAY DIFFRACTION11B1 - 31
12X-RAY DIFFRACTION12B32 - 132
13X-RAY DIFFRACTION13B133 - 155
14X-RAY DIFFRACTION14B156 - 178
15X-RAY DIFFRACTION15B179 - 212
16X-RAY DIFFRACTION16B213 - 260
17X-RAY DIFFRACTION17B261 - 280
18X-RAY DIFFRACTION18B281 - 304
19X-RAY DIFFRACTION19B305 - 374
20X-RAY DIFFRACTION20B375 - 384
21X-RAY DIFFRACTION21C1 - 7
22X-RAY DIFFRACTION22C8 - 55
23X-RAY DIFFRACTION23C56 - 115
24X-RAY DIFFRACTION24C116 - 153
25X-RAY DIFFRACTION25C154 - 178
26X-RAY DIFFRACTION26C179 - 255
27X-RAY DIFFRACTION27C256 - 283
28X-RAY DIFFRACTION28C284 - 301
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