Application of arginine as an efficient eluent in cation exchange chromatographic purification of a PEGylated peptide

Annathur, Gopinath V.; Buckley, John J.; Muthurania, Kevin; Ramasubramanyan, Natrajan

doi:10.1016/j.chroma.2010.04.019

Cited by 7 publications

(4 citation statements)

References 22 publications

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“…Historically, arginine has been used for improving the refolding efficiency of recombinant proteins in Escherichia coli that have been expressed into inclusion bodies (Arora and Khanna, 1996; Brinkmann et al, 1992). More recently, arginine has been used in protein purification applications to improve separation and recovery and to provide milder elution conditions for a variety of chromatographic resins (Annathur et al, 2010; Arakawa et al, 2004, 2007a; Ejima et al, 2005; Tsumoto et al, 2007). While the underlying mechanism is not well understood, arginine has been shown to improve protein stability and suppress aggregation by directly binding to proteins and by indirect methods such as altering water structure (Arakawa et al, 2007b; Kita et al, 1994; Lin and Timasheff, 1996; Schneider and Trout, 2009; Shukla and Trout, 2010; Shukla et al, 2011).…”

Section: Introductionmentioning

confidence: 99%

Mobile phase modifier effects in multimodal cation exchange chromatography

Holstein

Parimal

McCallum

et al. 2011

Biotech & Bioengineering

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This study examines protein adsorption behavior and the effects of mobile phase modifiers in multimodal chromatographic systems. Chromatography results with a diverse protein library indicate that multimodal and ion exchange resins have markedly different protein binding behavior and selectivity. NMR results corroborate the stronger binding observed for the multimodal system and provide insight into the structural basis for the observed binding behavior. Protein-binding affinity and selectivity in multimodal and ion exchange systems are then examined using a variety of mobile phase modifiers. Arginine and guanidine are found to have dramatic effects on protein adsorption, yielding changes in selectivity in both chromatographic systems. While sodium caprylate leads to slightly weaker chromatographic retention for most proteins, certain proteins exhibit significant losses in retention in both systems. The presence of a competitive binding mechanism between the multimodal ligand and sodium caprylate for binding to ubiquitin is confirmed using STD NMR. Polyol mobile phase modifiers are shown to result in increased retention for weakly bound proteins and decreased retention for strongly bound proteins, indicating that the overall retention behavior is determined by a balance between changes in electrostatic and hydrophobic interactions. This work provides an improved understanding of protein adsorption and mobile phase modifier effects in multimodal chromatographic systems and sets the stage for future work to develop more selective protein separation systems.

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Section: Introductionmentioning

confidence: 99%

Mobile phase modifier effects in multimodal cation exchange chromatography

Holstein

Parimal

McCallum

et al. 2011

Biotech & Bioengineering

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“…It has also been shown to induce unique selectivities for a library of proteins in ion‐exchange and multimodal systems . Arginine has also been widely used in ion‐exchange, hydrophobic, multimodal, and affinity chromatography …”

Section: Introductionmentioning

confidence: 99%

“…11 It has also been shown to induce unique selectivities for a library of proteins in ion-exchange and multimodal systems. 3 Arginine has also been widely used in ion-exchange, 4,12 hydrophobic, 4,13 multimodal, 8 and affinity chromatography. 14,15 Significant work has been done to explore the underlying mechanisms of these protein-guanidinium interactions, focused primarily on protein denaturation.…”

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confidence: 99%

Effect of guanidine and arginine on protein–ligand interactions in multimodal cation‐exchange chromatography

Parimal

Garde

Cramer

2017

Biotechnology Progress

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The addition of fluid phase modifiers provides significant opportunities for increasing the selectivity of multimodal chromatography. In order to optimize this selectivity, it is important to understand the fundamental interactions between proteins and these modifiers. To this end, molecular dynamics (MD) simulations were first performed to study the interactions of guanidine and arginine with three proteins. The simulation results showed that both guanidine and arginine interacted primarily with the negatively charged regions on the proteins and that these regions could be readily predicted using electrostatic potential maps. Protein surface characterization was then carried out using computationally efficient coarse-grained techniques for a broader set of proteins which exhibited interesting chromatographic retention behavior upon the addition of these modifiers. It was shown that proteins exhibiting an increased retention in the presence of guanidine possessed hydrophobic regions adjacent to negatively charged regions on their surfaces. In contrast, proteins which exhibited a decreased binding in the presence of guanidine did not have hydrophobic regions adjacent to negatively charged patches. These results indicated that the effect of guanidine could be described as a combination of competitive binding, charge neutralization and increased hydrophobic interactions for certain proteins. In contrast, arginine resulted in a significant decrease in protein retention times primarily due to competition for the resin and steric effects, with minimal accompanying increase in hydrophobic interactions. The approach presented in this paper which employs MD simulations to guide the application of coarse-grained approaches is expected to be extremely useful for methods development in downstream bioprocesses. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:435-447, 2017.

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“…There is a large body of literature examining the effects of mobile phase modifiers on protein retention behavior in traditional modes of chromatography. − There have also been a few recent papers on the effects of mobile phase modifiers in MM chromatography. Gao et al investigated the effects of mobile phase conditions, pH, and salt concentration on the adsorption behavior of bovine serum albumin on a MM cation exchange adsorbent developed for expanded bed adsorption. , We have recently carried out detailed studies into the effects of different mobile phase additives on protein adsorption and selectivity in MM chromatography. − …”

Section: Introductionmentioning

confidence: 99%

Effects of Urea on Selectivity and Protein–Ligand Interactions in Multimodal Cation Exchange Chromatography

et al. 2012

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Nuclear magnetic resonance (NMR) and molecular dynamics (MD) simulations were employed in concert with chromatography to provide insight into the effect of urea on protein-ligand interactions in multimodal (MM) chromatography. Chromatographic experiments with a protein library in ion exchange (IEX) and MM systems indicated that, while urea had a significant effect on protein retention and selectivity for a range of proteins in MM systems, the effects were much less pronounced in IEX. NMR titration experiments carried out with a multimodal ligand, and isotopically enriched human ubiquitin indicated that, while the ligand binding face of ubiquitin remained largely intact in the presence of urea, the strength of binding was decreased. MD simulations were carried out to provide further insight into the effect of urea on MM ligand binding. These results indicated that, while the overall ligand binding face of ubiquitin remained the same, there was a reduction in the occupancy of the MM ligand interaction region along with subtle changes in the residues involved in these interactions. This work demonstrates the effectiveness of urea in enhancing selectivity in MM chromatographic systems and also provides an in-depth analysis of how MM ligand-protein interactions are altered in the presence of this fluid phase modifier.

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Application of arginine as an efficient eluent in cation exchange chromatographic purification of a PEGylated peptide

Cited by 7 publications

References 22 publications

Mobile phase modifier effects in multimodal cation exchange chromatography

Mobile phase modifier effects in multimodal cation exchange chromatography

Effect of guanidine and arginine on protein–ligand interactions in multimodal cation‐exchange chromatography

Effects of Urea on Selectivity and Protein–Ligand Interactions in Multimodal Cation Exchange Chromatography

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