The plasmid for putidaredoxin expression (pPdX) was obtained from AddGene (plasmid 85084). PdX was purified and expressed as previously described.33 DEA- and PROLI-NONOates were purchased from Cayman Chemicals. DEA and PROLI-NONOate stocks were dissolved in 10 mM NaOH and quantified by measuring the absorbance at 250 (ε250 = 6500 M−1 cm−1) and 252 nm (ε = 8400 M−1 cm−1), respectively. General reagents and medium components were purchased from Fisher Scientific or VWR. Isopropyl β-d -1-thiogalactopyranoside (IPTG) and 5-amino-levulinic acid (5-ALA) were purchased from GoldBio. Water used for all solutions was of 18.2 MΩ cm resistivity from a Barnstead Nanopure system (Thermo Fisher Scientific). Solvents for LC-MS experiments were of at least HPLC grade and contained 0.3% (v/v) formic acid.
5 aminolevulinic acid 5 ala
Manufactured by GoldBio
5-aminolevulinic acid (5-ALA) is a naturally occurring compound that serves as a precursor in the biosynthesis of important biomolecules, such as heme and chlorophyll. It is a colorless, crystalline solid that is soluble in water and organic solvents.
Automatically generated - may contain errors
3 protocols using 5 aminolevulinic acid 5 ala
1
Purification and Quantification of PdX Redoxin
2
Reagent Preparation for Biochemical Assays
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Isopropyl β-ᴅ-1-thiogalactopyranoside (IPTG) and 5-aminolevulinic acid (5-ALA) were purchased from Gold Biotechnology. NNG was purchased from AAblocks. 2-NAE was purchased from Toronto Research Chemicals. General buffers and media components were purchased from Fisher Scientific or VWR. Stock dithionite concentrations were determined by UV–vis absorbance at 318 nm (ε318 = 8000 M−1cm−1). Water used for all solutions was of 18.2 MΩ·cm resistivity from a Barnstead Nanopure (Thermo Fisher Scientific). Solvents for LC–MS experiments were of at least HPLC grade and contained 0.1% vol/vol formic acid.
3
Purification and Quantification of Redox-Active Compounds
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The plasmid for putidaredoxin expression (pPdX) was obtained from AddGene (Plasmid #85084) and the recombinant protein purified and expressed as previously described. 53 DEA-and PROLI-NONOates were purchased from Cayman Chemicals. DEA and PROLI-NONOate stocks were dissolved in 10 mM NaOH and quantified by measuring the absorbance at 250 (ε250 = 6500 M -1 cm -1 ) or 252 nm (ε = 8400 M -1 cm -1 ), respectively. General reagents and media components were purchased from Fisher Scientific or VWR. Isopropyl β-D-1-thiogalactopyranoside (IPTG) and 5-aminolevulinic acid (5-ALA) were purchased from GoldBio. Water used for all solutions was of 18.2 MΩ•cm resistivity from a Barnstead Nanopure (Thermo Fisher Scientific). Solvents for LC-MS experiments were of at least HPLC grade and contained 0.3% v/v formic acid.
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