RSC Advances
·
2015
·Vol. 5
·期 82
·67093-67101
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IF 3.84
Yuping Wei,Jiandong Xu,Liang Zhang,Yankai Fu,Xia Xu
The huge demand for the most promising biopharmaceuticals, monoclonal antibodies (mAb), has resulted in a need for more efficient and less costly downstream purification processes for mAb. The currently used Protein A Agarose is unattractive due to toxic ligand leakage and high cost. In this study, three novel small peptide ligands, DWHW, CEWW and HEYW, were designed based on the interactions with human immunoglobulin G (IgG) using molecular simulations. The effects of pH, ionic strength and flow rate on the binding capacities to IgG were investigated. The static and dynamic binding capacities were determined. The dissociation constant (Kd) of the DWHW resin was 1.1 × 10−5 M. The binding capacity of the DWHW resin was 24.5 mg ml−1, which is comparable to Protein A Agarose. The DWHW resin was able to purify IgG from cMEM and CHO cell culture supernatants with a purity of more than 95% under elution conditions of pH 9 (50 mM sodium borate buffer) and pH 3 (50 mM glycine–HCl buffer). The results indicate that the small peptide ligands, especially DWHW, can offer a potential alternative for mAb purification.
研究领域:
生物化学
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生物工程
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制药工程
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DOI
高影响因子
JOURNAL OF CHEMICAL AND ENGINEERING DATA
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2019
·
IF 2.298
Hong-Yun Zhu,Sheng-Gang Chen,Qilei Zhang,Dong-Qiang Lin,Shan-Jing Yao
Alternative antibody-binding affinity ligands are expected to support robust time- and cost-effective antibody purification with rapid improvement of upstream productivity and demand on downstream process optimization. Five new tetrapeptide biomimetic affinity ligands (FYRH, HWRH, YHRI, HYRF, and FHRA) were screened by modeling the molecular interaction mechanism between tetrapeptide ligands and the Fc fragment of IgG with molecular dynamics simulation and the flexible docking method, and the corresponding five resins were prepared in this work to evaluate their adsorption characteristics of human immunoglobulin G (hIgG). The results showed that the binding capacities to hIgG of the five resins showed a strong pH-dependency, which was high at pH 7.0–9.0, and extremely low at pH 5.0–6.0, providing a benefit choice to separate hIgG. The dynamic binding capacities (DBC) at 10% breakthrough of five resins could reach up to 28–37 mg/mLgel, and Ac-FYRH-4FF resin showed the best adsorption performance. The elution test results with Ac-FYRH-4FF resin indicated that hIgG could be eluted mildly at pH 5.0 with a 96.5% recovery, meaning that this resin can be considered as a powerful candidate of alternative biomimetic affinity chromatographic resin.
研究领域:
生物化学
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蛋白质纯化
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高影响因子
BIOCHEMICAL ENGINEERING JOURNAL
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2016
·
IF 2.463
Rong-Zhu Wang,Dong-Qiang Lin,Wen-Ning Chu,Qi-Lei Zhang,Shan-Jing Yao
Peptide ligands have been developed as powerful alternatives to Protein A for antibody purification. In this study, a tetrapeptide library with two aromatic residues, one arginine and one aromatic/aliphatic residue (Aro-Aro-R-Aro/Ali) was constructed considering molecular interactions and applications. The library had 128 peptide ligands and was screened by flexible docking and molecular dynamics simulation. The results indicated that Ac-YFRH was the best ligand, and then was coupled onto an aminated argarose matrix. The results of isotherm adsorption showed that the binding capacity of human immunoglobulin G (IgG) was high at pH 7⿿8 and low at pH 4⿿5. The IgG adsorption also showed obvious salt-independence, and the addition of salt could reduce unspecific adsorption caused by electrostatic interaction. After the optimization of pH and salt addition, high IgG purity of 98.4% could be obtained with the recovery of 89.4% from BSA-containing feedstock. The purification of monoclonal antibody from CHO cell culture supernatant was tested, and the purity and recovery reached 98.0% and 79.5%, respectively. The results demonstrated that the new resin with the tetrapeptide YFRH ligand had high affinity and flexible operation window for IgG purification.
研究领域:
生物化学
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分子生物学
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高影响因子
BIOCHEMICAL ENGINEERING JOURNAL
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2021
·
IF 3.978
Yu-Ming Fang,Dong-Qiang Lin,Shan-Jing Yao
Short peptide resin has a great advantage in the separation and purification of antibodies due to its high selectivity and stable ligand binding ability. In this study, a peptide library containing 480 tetrapeptides was established with six critical residues (Phe, Tyr, Leu, Glu, Ile, Lys) of Protein A binding to Fc part of the antibody and one Histidine . Then the library was screened by LibDock and five tetrapeptide ligands with high LibDock scores were selected. Subsequently, the electrostatic interaction energies and van der Waals interaction energies between the screened ligands and the Fc-A fragment of IgG were further calculated by molecular dynamics simulation to research the binding ability between ligands and target proteins. The simulation results showed that YEHF was the optimal ligand with higher van der Waals interaction energies. Then, four tetrapeptides (YFLH, FYHI, FYKH and YEHF) were selected as ligands for preparing adsorption resins, and the resins were used to bind hIgG and BSA by static adsorption experiments. The results showed that Ac-YEHF-4FF resin had higher hIgG binding ability and lower BSA binding ability, also indicating that YEHF was the optimal screened ligand. Therefore, molecular simulation was verified as a well use technology for antibody separation and purification.
研究领域:
生物化学
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分子生物学
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DOI
高影响因子
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
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2019
·
IF 2.813
Jiawen Li,Dongqiang Lin,Shanjing Yao,N.K.H. Slater,Qilei Zhang
Traditional chromatographic techniques used in downstream processes of biomolecule manufacturing are often time-consuming and expensive. In this study, a cost-effective microporous micro-capillary film (MMCF) composed of ethylenevinyl alcohol (EVOH) was evaluated for its potential application in immunoadsorption with high process efficiency. A peptide ligand Ac-Phe-Tyr-His-Glu (Ac-FYHE) was immobilized on the inner surface of MMCF for selective binding of human immunoglobulin (hIgG). The porous structure and chemical properties of the prepared MMCF were confirmed by scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR). hIgG (2 mg/ml) adsorption studies demonstrated that the binding process followed a Langmuir isotherm with equilibrium adsorption capacities of 9.31 and 3.47 mg/ml adsorbent under static and dynamic conditions, respectively. Moreover, the membrane showed good flowrate tolerance when studied under flowrates of 0.5 ml/min to 10 ml/min. hIgG purity was 88.2% when obtained from an hIgG (2 mg/ml) and HSA (8 mg/ml) mixture and the purity remained over 80.0% when hIgG concentrations increased in the mixtures. Moreover, purity of 82.3% was achieved when removing hIgG directly from human serum. The MMCF-Ac-FYHE affinity column is expected to selectively remove hIgG from blood for the treatment of autoimmune diseases with high efficiency and cost effectiveness.
研究领域:
生物工程
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医学工程
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产品关联
DOI
PubMed
高影响因子
EUROPEAN JOURNAL OF PHARMACEUTICS AND BIOPHARMACEUTICS
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2014
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IF 4.245
Tingting Fan,Chunhui Chen,Han Guo,Juan Xu,Jian Zhang,Xi Zhu,Yang Yang,Zhou Zhou,Lian Li,Yuan Huang
Designing feasible and effective peptide ligand modified solid lipid nanoparticles (SLNs) to improve oral bioavailability of protein drugs and evaluating the influence of mucus remains important. In the present work, two kinds of peptide ligand modified SLNs loaded with salmon calcitonin (sCT), namely, sCT CSK-SLNs and sCT IRQ-SLNs, were prepared by coupling the peptide ligand CSKSSDYQC (CSK) which was reported to show affinity with goblet cells, or IRQRRRR (IRQ), a cell penetrating peptide, to polyoxyethylene (40) stearate (SA-PEG 2000 ). Compared with unmodified SLNs, CSK or IRQ modified SLNs with better drug protection ability could facilitate the internalization of drug on Caco-2/HT29-MTX co-cultured cells and permeation in excised rat duodenum mucosa. The internalization mechanism of two kinds of peptide ligand modified SLNs was mainly active transport via both clathrin- and caveolae-dependent endocytosis. Although mucus was an impediment to the transport of SLNs, the peptide ligand modified SLNs still showed improved drug absorption. The absolute bioavailability of sCT CSK-SLNs (12.41 ± 3.65%) and sCT IRQ-SLNs (10.05 ± 5.10%) raised to 2.45-fold and 1.98-fold compared with unmodified SLNs (5.07 ± 0.54%), implying the feasibility and effectiveness of CSK and IRQ peptide modification for the enhancement of the oral bioavailability of protein drugs. In summary, the nanoparticles modified with CSK or IRQ peptide ligand could be the potential carriers for the transport of protein drugs across intestinal barriers.
研究领域:
药剂学
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生物医学工程
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分子生物学
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DOI
PubMed
高影响因子
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
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2022
·
IF 3.318
Yu-Ming Fang,Qi-Lei Zhang,Dong-Qiang Lin,Shan-Jing Yao
Short peptide biomimetic chromatography technology as a developing protein separation technology has huge potential for antibody purification. In this study, four tetrapeptide ligands (Ac-FYKH, Ac-YEHF, Ac-YFLH and Ac-FYHI) with high potential binding ability to antibody were selected for the optimal ligand to antibody purification. The results showed that Ac-YEHF-4FF resin had higher binding capacity and selectivity for hIgG among the four resins. And at pH 7.0 and 0.3 ml/min, the highest Q 10%-hIgG of Ac-YEHF-4FF resin was 26.2 mg/ml resin while its Q 10%-BSA was just 2.2 mg/ml resin. Further, Ac-YEHF-4FF resin was used to purify protein mixtures. By binding at pH 7.0 and being eluted at pH 5.0 and pH 4.0, Ac-YEHF-4FF resin was well used to separate hIgG from BSA containing feedstock, HSA containing feedstock and human serum with the purity and yield both more than 95 %. And the screened resin could also separate mAb from CHO cell culture supernatant with purity 94.3 % and yield 97.5 %. The adsorption and separation results of Ac-YEHF-4FF resin indicated that the goal of getting the efficacy of critical residues from protein A to biomimetic its structure and function could be achieved, which had great significance to the establishment and improvement of tetrapeptide biomimetic chromatography, and also provided a new method for the field of antibody separation and purification.
研究领域:
生物化学
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蛋白质工程
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分离技术
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DOI
PubMed
高影响因子
JOURNAL OF CHROMATOGRAPHY A
·
2019
·
IF 3.858
Yu-Ming Fang,Sheng-Gang Chen,Dong-Qiang Lin,Shan-Jing Yao
Biomimetic affinity chromatography with short peptide ligands is a developing technology, which has great potential for antibody separation and purification. In this study, a tetrapeptide library with critical residues of natural ligands to hIgG was constructed and a novel tetrapeptide ligand (Ac-FYHE) with high LibDock scores was selected by molecular docking. Then, Ac-FYHE ligand was linked to agarose bead to prepare a new chromatography resin. The properties of antibody adsorption were measured and evaluated by static/dynamic adsorption. It was found that the resin with ligand Ac-FYHE has high binding capacity and selectivity for hIgG. The results showed the Q m-hIgG of Ac-FYHE-4FF resin was 87.9 mg/g resin while the Q m-BSA of this resin was only 16.5 mg/g resin at pH 7.0. Moreover, at pH 7.0, Q 10% of Ac-FYHE-4FF resin was 24.1 mg/mL for hIgG but just 2.1 mg/mL for BSA, which presented high selectivity of the screened resin at pH 7.0. Subsequently, the adsorption and separation properties of the Ac-FYHE-4FF resin were further investigated. As a result, with the addition of 0.5 M NaCl , Q m decreased by less than 20% but Q m decreased by 70% with the addition of 50% (v/v) ethylene glycol , which indicated that hydrophobic interaction would be the driving force for the binding between resin and hIgG. Besides, pH 7.5 and pH 4.5 could be the optimal loading and elution condition for hIgG, respectively. Finally, the Ac-FYHE-4FF resin was applied to separate mAb or/and hIgG from BSA containing feedstock, CHO cell culture supernatant and human serum, and the purity and recovery were both more than 90% with only one-step separation. The results indicate that the Ac-FYHE-4FF resin developed in this work would be promising for antibody separation and purification.
研究领域:
生物化学
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生物工程
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制药工程
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DOI
PubMed
高影响因子
Peptide Science
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2022
·Vol. 115
·期 2
·e24302
·
IF 2.592
Anqi Wang,Kai Yue,Yiang Wei,Weishen Zhong,Genpei Zhang
The structural change of receptor protein at high temperatures is one of the factors affecting the targeting ability of ligand-installed nanocarriers for combined therapy of hyperthermia and drug delivery. In this study, the binding behaviors and mechanisms of integrin αvβ3 receptor and the arginine-glycine-aspartic acid (RGD) peptide ligand at high temperatures were investigated both theoretically and experimentally. The structural parameters of integrin αvβ3 at different temperatures and the interaction forces between the RGD peptide and integrin αvβ3 at different binding sites were calculated by molecular dynamics simulation. Fourier transform infrared spectroscopy, energy dispersive spectroscopy, ultraviolet–visible absorption spectroscopy, and atomic force microscopy were used to analyze the structural changes of integrin αvβ3 and to measure the ligand-receptor interaction. Results show that the number of hydrogen bonds decreased and the secondary structure of integrin αvβ3 changed with the increase in temperature, indicating the denaturation of integrin αvβ3. The structural stability of the integrin αv subunit was better than that of the integrin β3 subunit at high temperatures. The interaction between the RGD peptide and integrin αvβ3 weakened as the temperature increased because the structure of the integrin αvβ3 binding site became more flexible and the corresponding calcium ions were shed from the binding site. The strongest interaction force was exhibited at the binding site of the integrin β3 subunit at 310 K while it was found at the binding site of the integrin αv subunit at higher temperatures, owing to the smaller structure deformation of the integrin αv subunit. Graphical
研究领域:
生物物理学
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分子生物学
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纳米医学
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DOI
高影响因子
MICROCHEMICAL JOURNAL
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2024
·
IF 4.9
Xintong Li,Jiale Cui,Jin Li,Yulian Wang,Guoqing Shi
Gastrin 17 (G-17) is a major biomarker for the diagnosis of gastric cancer. As the serum concentrations of gastrin are low, highly sensitive detection methods are needed. Previous studies have shown that oriented immobilization of detection antibodies can improve the sensitivity of lateral flow immunoassay (LFIA) methods. Herein, the antibody binding capacity of four Fc-specific peptides reported in different studies was evaluated, and two peptides with better binding potency were further modified with cysteine or the peptide GAC at their C-terminus. The peptide FYWHCLDEGAC exhibited the highest antibody binding capacity and was applied to directly immobilize the anti-G-17 antibody on microspheres with carboxyl-functionalized surfaces and europium-based time-resolved fluorescence. An LFIA method was developed to detect G-17 in human serum, and the limit of detection (LOD) of this method for G-17 was 0.08 pmol L -1 , which was 10 times less than that of the nonoriented immobilization method. The working range of this method is 0.27 ∼ 600 pmol L -1 , with a recovery rate of 85.2 % −106.7 % and a CV value less than 12.4 %. This method is easy to use, and the total detection process takes less than 20 min. The test results for the actual serum samples were consistent with the commercial kit results (R 2 > 0.95), demonstrating that the method can be utilized for point-of-care testing of G-17. This approach to orientedly immobilize antibodies on nanomaterials can be used to develop sensitive LFIA methods for various molecules.
研究领域:
生物医学工程
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分析化学
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癌症研究
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分子生物学
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DOI
高影响因子
Acta Pharmaceutica Sinica B
·
2023
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IF 14.5
Jiawen Yang,Aixuan Zhou,Minyi Li,Qiaoxian He,Jingwei Zhou,Jacques Crommen,Wentao Wang,Zhengjin Jiang,Qiqin Wang
Due to low immobilized ligand density, limited binding capacity, and severe interference from serum proteins, developing ideal peptide-based biomaterials for precise recognition and in vivo analysis of biopharmaceuticals remains a huge challenge. In this study, mimotope peptide modified pompon mum-like biomimetic magnetic microparticles (MMPs, 3.8 μm) that mimic the specific functionalities of CD20 on malignant B cells were developed for the first time. Benefit from the numerous ligand binding sites (Ni 2+ ) on the pompon mum-like MMPs, these novel materials achieved ≥10 times higher peptide ligand densities (>2300 mg/g) and antibody binding capacities (1380 mg/g) compared to previous reported biomaterials. Leveraging the high specificity of the mimotope peptide, rituximab can be precisely recognized and enriched from cell culture media or serum samples. We also established an LC‒MS/MS method using the MMPs for tracking rituximab biotransformation in patient serum. Intriguingly, deamidation of Asn55 and Asn33, as well as oxidation of Met81 and Met34 were observed at the key complementarity determining regions of rituximab, which could potentially influence antibody function and require careful monitoring. Overall, these versatile biomimetic MMPs demonstrate superior recognition and enrichment capabilities for target antibodies, offering interesting possibilities for biotransformation analysis of biopharmaceuticals in patient serum.
研究领域:
生物技术
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药学
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分析化学
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生物化学
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DOI
PubMed
高影响因子
ACS Applied Materials & Interfaces
·
2017
·
IF 7.504
Danni Ran,Jiani Mao,Changyou Zhan,Cao Xie,Huitong Ruan,Man Ying,Jianfen Zhou,Wan-Liang Lu,Weiyue Lu
Compared to that of other tumors, various barriers, such as the blood–brain barrier (BBB), enzymatic barriers, and the blood–brain tumor barrier, severely impede the successful treatment of gliomas. Peptide ligands were frequently used as targeting moieties to mediate brain tumor-targeted drug delivery. LWSW (SYPGWSW) is a recently reported quorum-sensing (QS) peptide that is able to efficiently cross the BBB. Even though linear LWSW traverses the BBB in vitro, its in vivo targeting ability has been greatly impaired due to proteolysis. Here, we developed a stable peptide, DWSW (DWDSDWDGDPDYDS), using the retro-inverso isomerization technique to achieve an enhanced antiglioma effect. In vitro studies have demonstrated that both the LWSW and DWSW peptides possessed excellent tumor-homing properties and barrier-penetration abilities, whereas DWSW exhibited exceptional stability in serum and maintained its targeting ability after serum preincubation. In vivo, DWSW-modified probes and micelles accumulated more efficiently in the glioma region in comparison with LWSW-modified probes and micelles because of full resistance to proteolysis in blood circulation. As expected, DWSW-modified paclitaxel (PTX)-loaded micelles (DWSW Micelle/PTX) exhibited the longest median survival time among glioma-bearing nude mice. Our results suggested that the QS peptide appears to be a promising targeting moiety, with potential applications in glioma-targeted drug delivery.
研究领域:
药学科学
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神经科学
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癌症研究
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药物递送系统
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生物医学工程
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DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）