7QSX

Non-obligately L8S8-complex forming RubisCO derived from ancestral sequence reconstruction and rational engineering in L8 complex

Experimental procedure

Experimental method	SINGLE WAVELENGTH
Source type	SYNCHROTRON
Source details	ESRF BEAMLINE ID23-2
Synchrotron site	ESRF
Beamline	ID23-2
Temperature [K]	100
Detector technology	PIXEL
Collection date	2021-07-09
Detector	DECTRIS PILATUS3 2M
Wavelength(s)	0.873128
Spacegroup name	P 1 21 1
Unit cell lengths	113.599, 159.197, 127.910
Unit cell angles	90.00, 107.90, 90.00

Refinement procedure

Resolution	29.889 - 2.700
R-factor	0.2189
Rwork	0.218
R-free	0.26030
Structure solution method	MOLECULAR REPLACEMENT
Starting model (for MR)	6ura
Data reduction software	XDS (20210323)
Data scaling software	SCALA (3.3.22)
Phasing software	PHENIX (1.18.2_3874)
Refinement software	PHENIX (1.18.2_3874)

Data quality characteristics

	Overall	Inner shell	Outer shell
Low resolution limit [Å]	29.889	29.889	2.850
High resolution limit [Å]	2.700	8.540	2.700
Rmerge		0.092	0.821
Rmeas	0.298	0.101	0.895
Rpim	0.119	0.043	0.353
Total number of observations	708705	17748	107444
Number of reflections	118081	3468	17291
<I/σ(I)>	5.8	11.3	2.2
Completeness [%]	99.6	91.2	100
Redundancy	6	5.1	6.2

Crystallization Conditions

crystal ID	method	pH	temperature	details
1	VAPOR DIFFUSION, SITTING DROP	6.5	288	Purified enzyme (5 mg/mL) in 25 mM Tricine-NaOH, 75 mM NaCl, pH 8.0 was incubated at 3% (v/v) CO2 in air and 30 degrees C for 1 h in the presence of 0.2 mM CABP and 3.2 mM MgCl2. The enzyme was then mixed in a 1:1 ratio with 0.1 M ammonium sulfate, 0.3 M sodium formate, 0.1 M sodium cacodylate , 3% (w/v) gamma-PGA, and 2% (w/v) PEG 3350, pH 6.5. Before flash freezing the crystals in liquid nitrogen PEG200 was added to the mother liquor as cryoprotectant to a final concentration of 40 % (w/v).