Stereospecific interactions between histidine and monoclonal antibodies

Abstract

Histidine is a frequently used buffer in the final formulation of many commercialized monoclonal antibodies (mAbs), with histidine helping to stabilize the antibody during storage in addition to its buffering function. The objective of this study was to examine the stereospecificity of any histidine-antibody interactions using a combination of experimental studies and molecular dynamics simulations. Isothermal titration calorimetry provided evidence of weak stereospecific interactions, with the antibody showing approximately two to four additional interaction sites for D-versus L-histidine. The greater interactions with D-histidine were confirmed by measurements of the net protein charge using electrophoretic light scattering. The reduction in the net negative charge of the antibody in D-histidine led to significantly different behavior during diafiltration due to Donnan exclusion effects. Molecular dynamics simulations corroborated the presence of additional D-histidine interaction sites. These results provide the first demonstration of weak stereospecific interactions between L-and D-histidine and a mAb and the implications of these interactions for antibody formulation. K E Y W O R D S buffer interactions, formulation, histidine, monoclonal antibody, stereospecificity

Membrane fouling by lysozyme: Effect of local interaction

“…For consistency, CHARMM force field parameters were used for both water and ions. The SHAKE algorithm was applied to all bonds involving hydrogen 27,60 . To stabilize the system, energy minimization was performed first with a canonical ensemble (NVT) run at 30 K for 100 ps, followed by another NVT run to bring the system up to room temperature (298 K), after which the system was relaxed at 298 K by another 500 ps NVT run.…”

“…All MD simulations were run using large‐scale atomic/molecular massively parallel simulator (LAMMPS), 58 with visualization and post‐processing facilitated by the visual molecular dynamics (VMD) software 59 . Pair‐wise non‐bonded interactions in LAMMPS were set with “lj/charmm/coul/long” style with an outer cut‐off distance of 10 Å, which is in line with other MD studies on proteins 36,60 . The PPPM Ewald method was used for computing long‐range electrostatics beyond this cut‐off in reciprocal space.…”

Process analytical technology for on‐line monitoring of quality attributes during single‐use ultrafiltration/diafiltration

“…The three SEC‐based methods were developed as a PAT tool kit, each designed to be fit‐for‐purpose and relatively simple, which aligns with the need for robustness in PAT, and to maximize the value of the data generated. Each method was optimized to obtain mAb purity and excipient concentrations over the wide range of protein concentrations and buffer matrix conditions that a UF/DF process covers (Baek et al, 2019, 2019; Le et al, 2014; Powers et al, 2019). These PAT methods can accurately quantitate mAb purity and the concentration of the excipients, histidine and arginine without the need of chemical tag.…”

Molecular dynamics study on membrane fouling by oppositely charged proteins

“…The simulations were run in Large‐scale Atomic/Molecular Massively Parallel Simulator (LAMMPS) 61 . All light bonds (i.e., covalent bonds involving hydrogen atoms) were constrained by the SHAKE algorithm, which is commonly implemented to increase stability and save computation cost 62 . Long‐range electrostatic interactions were computed by the PPPM Ewald method in the reciprocal space.…”

“…Long‐range electrostatic interactions were computed by the PPPM Ewald method in the reciprocal space. The Lennard–Jones (LJ) non‐bonded interaction was switched off by applying inner‐cutoffs and outer‐cutoffs of 8 and 10 , respectively, which are common thresholds 31,62 . Energy minimization was performed first, followed by a canonical ensemble (NVT) run that increases system temperature from 30 to 298 K. The system was subsequently relaxed at 298 K with a 500 ps NVT run, followed by an equilibration NPT (isothermal–isobaric ensemble) run for 1 ns.…”