Structure-based design and application of a nucleotide coenzyme mimetic ligand: Application to the affinity purification of nucleotide dependent enzymes

Reduce, Reuse and Recycle in Protein Chromatography: Development of an Affinity Adsorbent from Waste Paper and Its Application for the Purification of Proteases from Fish By-Products

“…The adsorption kinetics of BSA and LYS onto CB3GA-Cellulose-2 adsorbent is shown in Figure 6. The results indicate that the overall rate of the BSA and LYS adsorption process is most likely to be controlled by the chemisorption process [32][33][34][35]. The best fit values of h, q e and k 2 for the pseudo-second-order models are shown in Table 2.…”

“…The ratio K 1 /K 2 equals the equilibrium dissociation constant (K D ) of the enzyme-ligand complex. The Langmuir isotherm (Equation (2)) has been widely used to describe protein adsorption onto a wide range of affinity adsorbents [29][30][31][32]:…”

“…Adsorption kinetics studies were carried out as previously described [29,30,32], with minor modifications: in a total volume of 4 mL (10 mM potassium phosphate buffer pH 7.0, protein (BSA or LYS, 5 mg total protein), in 10 mM potassium phosphate buffer pH 7.0, was mixed with the CB3GA-Cellulose-2 affinity adsorbent (100 mg moist wet gel, 9.3 mg dry mass). The suspension was shaken at 4 • C. Adsorption velocity was monitored by periodically removing samples from the suspensions, which were subsequently centrifuged (10,000× g, 30 s), and the amount of total protein concentration in the supernatant was determined by the Bradford method [28].…”

“…Biomolecules 2020, 10, x FOR PEER REVIEW 9 of 13 controlled by the chemisorption process [32][33][34][35]. The best fit values of h, qe and k2 for the pseudosecond-order models are shown in Table 2.…”

“…It is well established that the pH of the binding and elution buffer is one of the most crucial parameters in dye-ligand affinity chromatography as it can affect dramatically the chromatographic behavior, such as affinity, selectivity and recovery of the target protein [29,30,41]. The results showed that at acidic pH values the binding capacity is enhanced, compared to that at controlled by the chemisorption process [32][33][34][35]. The best fit values of h, qe and k2 for the pseudosecond-order models are shown in Table 2.…”