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26 protocols using «707 sq 250m»

1

X-band EPR Analysis of MagR Proteins

2024
X-band (9.6 GHz) EPR spectra of the MagR proteins were recorded using an EMX plus 10/12 spectrometer (Bruker, USA), equipped with an Oxford ESR-910 liquid helium cryostat. Briefly, 1 mmol/L oxidized proteins (as-purified) in buffer E (20 mmol/L Tris, 150 mmol/L NaCl, 5 mmol/L d-desthiobiotin, pH8.0) were mixed in a total volume of 200 μL with 50 μL of glycerol, respectively. The 1 mmol/L reduced proteins were obtained by adding 2 μL of Na2S2O4 (1 mol/L) to the protein samples. The protein samples were then transferred into a 4 mm diameter quartz EPR tube (Wilmad 707-SQ-250 M, USA) and frozen in liquid nitrogen. The EPR signals of the oxidized and reduced proteins were recorded at different temperatures (10, 25, 45, and 60 K). The EPR conditions were: microwave frequency, 9.4 GHz; microwave power, 2 mW; modulation frequency, 100 kHz; modulation amplitude, 2 G; receive gain, 1.0×104.
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2

EPR Spectroscopy of Iron-Sulfur Proteins

2024
The cells expressing clMagRWT, clMagR3M, ecIscA, EntB, FepA or none, were harvested and resuspended in LB medium containing 20 mM deferoxamine and incubated at 37°C for anther 15 min as described by the Imlay's group in 2002 [35 (link)]. Then, the cells were washed with 10 mM diethylenetriaminepentaacetic acid once, and 20 mM cold Tris-HCl (pH = 7.4) twice and resuspended in 20 mM cold Tris-HCl (pH = 7.4). 200 μL of the above samples were mixed with 50 μL of glycerol and transferred to an EPR tube (Wilmad 707-SQ-250 M, USA) and frozen in liquid nitrogen until EPR measurements. The EPR signals were monitored at different temperatures (10 K) with a microwave frequency of 9.40 GHz, a microwave power of 2 mW, a modulation amplitude of 2.0 G and a receive gain of 1.0 × 104.
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3

EPR Spectroscopy of Oxidized and Reduced Proteins

2024
X-band EPR spectra were recorded at various temperatures (10 K, 25 K, 45 K, and 60 K) using an EMX plus 10/12 Spectrometer (Bruker, Germany) equipped with an Oxford ESR910 Liquid Helium cryostat. The oxidized EPR samples were prepared by mixing 200 μL of 1 mmol/L purified protein with 50 μL of glycerol, while the reduced samples were obtained by adding sodium dithionite to a final concentration of 10 mmol/L to the above protein solutions. The protein samples were transferred to quartz EPR tubes (Wilmad 707-SQ-250 M, France) and frozen in liquid nitrogen. The EPR signals were collected at a microwave frequency of 9.40 GHz, microwave power of 2 mW, modulation amplitude of 2.0 G, receive gain of 1.0×104, and sweep time of 25.60 s.
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4

X-band EPR Spectroscopy of Oxidized Proteins

2024
X-band (9.6 GHz) electron paramagnetic resonance (EPR) spectra were recorded using an EMX plus 10/12 spectrometer (Bruker, Billerica, MA) equipped with an Oxford ESR-910 liquid helium cryostat. For oxidized protein samples, 50 µL of glycerol was added to 200 µL of 1 mmol/L sample. The protein samples were then injected into quartz EPR tubes (Wilmad 707-SQ-250 M, USA) and frozen with liquid nitrogen. The EPR signals were collected at different temperatures (10 K, 25 K, 45 K, and 60 K) under a microwave frequency of 9.40 GHz, microwave power of 2 mW, modulation amplitude of 2.0 G, and receive gain of 1.0×104.
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5

In situ EPR Analysis of eCO2RR

2024
Time-dependent continuous-wave (CW) EPR measurements at X-band frequencies (9.4 GHz) were performed using a Bruker EMX Micro EPR spectrometer. DMPO was used as a spin trap. The electrochemical cell used for in situ EPR testing is the same as that used for the in situ XAS setup. All electrochemistry procedures are identical, except that the electrochemical test was performed in a CO2-saturated 0.1 M KHCO3 electrolyte (pH = 6.8 ± 0.3) containing 10 mM of DMPO. During eCO2RR, 10 μL of samples were taken from the electrochemical cell and transferred into 1 mm ID, 2 mm OD EPR tube (712-SQ-100M Wilmad LabGlass). For EPR measurements, these tubes were placed into 3 mm ID 4 mm OD EPR tubes (707-SQ-250M Wilmad LabGlass). For all the measurements, the microwave power was set to 1.262 mW (22 dB microwave power attenuation) and modulation amplitude was 2 G at a modulation frequency of 100 kHz. EPR spectra were simulated using EasySpin software42 (link).
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