Butyl sepharose ff

Recombinant histidine-tagged apoAequorin (198 amino acids, Mw = 22,512.1) was expressed into the periplasmic space of E. coli cells using a piP-His-HE expression vector and purified, as previously described [19 (link)]. Coelenteramide (CTMD) was chemically synthesized according to a previously described procedure [19 (link)]. The following chemicals were obtained from commercial sources: coelenterazine (JNC. Co., Tokyo, Japan); 2-mercaptoethanol, dithiothreitol (DTT), dimethyl sulfoxide (DMSO), ethylenediaminetetraacetic acid disodium salt (EDTA·2Na), and imidazole (Wako Pure Chemicals, Osaka, Japan); Butyl-Sepharose FF (GE Healthcare, Piscataway, NJ, USA). All other chemicals were of the highest commercial grade available.

DEAE-Sepharose ff, Superdex-75, Butyl Sepharose ff, were purchased from GE Healthcare Life Sciences. Phosphoglucose isomerase from rabbit muscle, phosphomannose isomerase from E. coli, glucose 6-phosphate dehydrogenase from baker's yeast (S. cerevisiae), phosphoglucomutase from rabbit muscle, α-D-Glucose 1,6-bisphosphate potassium salt hydrate, α-D-Glucose 1-phosphate disodium salt hydrate, α-D(+)Mannose 1-phosphate sodium salt hydrate, D-Glucose 6-phosphate, D-Mannose 6-phosphate, β-Nicotinamide adenine dinucleotide phosphate sodium salt, were purchased from Sigma-Aldrich. Sypro Orange was from Invitrogen Molecular Probes. Sodium orthovanadate was from Acros Organics. Creatine phosphate disodium salt tetrahydrate was from Alfa Aesar. Clodronate ((Dichloro-phosphono-methyl)phosphonic acid) and neridronate ((6-Amino-1-hydroxyhexane-1,1-diyl)bis(phosphonic acid) were from ABIOGEN-PHARMA.
Mannose 1,6-bisphosphate was synthesized as described [18 (link)] and purified on an AG1x8 hydroxide form column by running a step gradient 0–1 M NaCl. Fractions were analyzed by ¹H- and ³¹P-NMR spectroscopy. An internal standard (Trimethylsilylpropanoic acid) was used in order to measure the concentration. All the other reagents were of analytical grade. The ORF encoding QDK-PMM1 was obtained by de novo gene synthesis and was purchased by GeneCust, Luxemburg; it is available upon request.