Kanamycin sulfate

Salts and other ingredients for buffers and
bacteria culture media,
dopamine hydrochloride, fetal bovine serum (FBS), l-glutamine,
sodium l-(+)-lactate, kanamycin sulfate, β-d-1-thiogalactopyranoside (IPTG), and phosphate-buffered saline (PBS)
were obtained from Sigma-Aldrich (St. Louis, MO). Alginic acid sodium
salt from brown algae (ref 71238), poly-l-lysine hydrochloride
(MW 15 000–30 000 Da), and low-molecular-weight
chitosan (ref 448869) were also supplied by Sigma-Aldrich. Chemically
competent E. coli NEB5α cells
were purchased from New England Biolabs (NEB, MA).
M9 culture
medium was prepared with 33.7 mM Na₂HPO₄, 22
mM KH₂PO₄, 8.55 mM NaCl, and 9.35
mM NH₄Cl as 10× stock and autoclaved. It was supplemented
with 0.4% d-glucose (or glycerol), 1 mM MgSO₄,
0.3 mM CaCl₂, and 1 mg/L thiamine from stocks that were
prepared separately and filter-sterilized. Krebs–Ringer N-(2-hydroxyethyl)piperazine-N′-ethanesulfonic
acid (HEPES) (KRH buffer) buffer was prepared with 20 mM HEPES, 135
mM NaCl, 5 mM KCl, 0.4 mM K₂HPO₄, adjusted to
pH 7.4, autoclaved, and supplemented with 1 mM MgSO₄, and
1 mM CaCl₂ from stocks prepared separately and filter-sterilized.
Incubation buffer (buffer A) consisted of 10 mM HEPES, 150 mM NaCl,
20 mM CaCl₂, and disruption buffer (buffer B) of 0.1 M
ethylenediaminetetraacetic acid (EDTA) and 0.2 M potassium citrate;
both were filter-sterilized.

Proli-NONOate was from Cayman chemicals, UK. Sodium phosphate (sodium dihydrogen orthophosphate and disodium hydrogen orthophosphate), sodium chloride, sodium tetraborate, sodium imidazole, kanamycin sulfate, and sodium chloride were purchased from Sigma-Aldrich, Poole, UK. Luria Bertani media was purchased from Melford laboratories Ltd., Ipswich, UK. Isopropyl B-D-1-thiogalactopyranoside and aminoleuvelic acid were obtained from Molekula Ltd., Darlington, UK.

All chemicals and reagents used in this study were of the highest purity grade. LB medium, kanamycin sulfate, isopropyl-β-D-thiogalactopyranoside (IPTG), nickel-nitrilotriacetic acid resin (Ni-NTA), imidazole, 4-nitrophenyl-β-D-glucopyranoside (pNPG), 1-anilinonaphthalene-8-sulfonate (ANS), tannic acid (TAN, molecular weight of 1,701.2 g/mol) and Triton X-100 (approximate molecular weight of 625 g/mol) were purchased from Sigma–Aldrich.

Staphylococcus epidermidis RP62A was obtained from American Type Culture Collection (ATCC 35984), and S. epidermidis clinical isolates were collected from the Department of Laboratory Medicine, Affiliated Gulou Hospital, Medical College of Nanjing University. Staphylococcal strains were cultured in tryptic soy broth (TSB; Oxoid Ltd., Basingstoke, United Kingdom), and Escherichia coli strains DH5α and BL21(DE3) were cultured in Luria-Bertani (LB) broth (1% tryptone, 0.5% yeast extract, 1% NaCl). Kanamycin sulfate was obtained from Sigma Aldrich (St. Louis, MO, United States).

Hesperetin (HST), kanamycin sulfate, chloramphenicol, D-sorbitol, β-mercaptoethanol (BME), isopropyl β-D-1-thiogalactopyranoside (IPTG), ethylenediamine tetraacetic acid (EDTA), dimethyl sulfoxide (DMSO), deuterated DMSO-d₆, deuterium oxide (²H₂O and D₂O) ribonuclease A, Tris-HCl, NaCl, ZnSO₄, (NH₄)₂SO₄, K₂HPO₄, and KH₂PO₄ were purchased from Sigma-Aldrich (Saint Louis, MO, USA); 1,4-dithiothreitol (DTT) from InLab; and Amicon Ultra-15 centrifugal filter (MWCO: 3.0 kDa) from Millipore. Hesperidin (HSD) was produced, purified [40 ] and kindly provided by Prof. Dr. Ljubica Tasic from University of Campinas (UNICAMP), Brazil.

Dichlorophenolindophenol (DCPIP) and potassium ferricyanide were from Fluka (Buchs, Switzerland). L-Arginine, ampicillin, kanamycin sulfate, chloramphenicol, cytochrome c (horse heart), isopropyl β-D-1-thiogalactopyranoside (dioxane-free), thiamine, glucose 6-phosphate, glucose 6-phosphate dehydrogenase, NADP⁺ and NADPH were obtained from Sigma–Aldrich (St. Louis, MO, United States). Phenylmethanesulfonyl fluoride was purchased from Gerbu Biotechnik GmbH (Heidelberg, Germany). Bacto agar, Bacto peptone and Bacto tryptone were obtained from BD Biosciences (San Jose, CA, United States). Bacto yeast extract was obtained from Formedium (Norwich, England). A polyclonal antibody from rabbit serum raised against recombinant human CPR obtained from Genetex (Irvine, CA, United States) was used for immunodetection of the membrane-bound CPR, while the respective antibody used the in the case of the soluble form of the enzyme was obtained from Thermofisher Scientific (Waltham, MA, United States).