Comparison of three activated agaroses for use in affinity chromatography: Effects on coupling performance and ligand leakage

Sommeren, A.P.G. van; Machielsen, P.A.G.M.; Gribnau, T.C.J.

doi:10.1016/0021-9673(93)83084-6

Cited by 29 publications

(8 citation statements)

References 16 publications

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“…Step e: L-Arg Affinity Chromatography-L-Arg-Sepharose was prepared from NHS-activated Sepharose 4 Fast Flow (Amersham Pharmacia Biotech) as described (35) including the following changes. In the coupling reaction, a solution of 500 mM L-Arg/HCl, 200 mM H 3 BO 3 , 500 mM NaCl, pH 8.2, was used.…”

Section: Protein Purificationmentioning

confidence: 99%

Structural and Functional Characterization of the Zn(II) Site in Dimethylargininase-1 (DDAH-1) from Bovine Brain

Knipp¹,

Charnock²,

Garner³

et al. 2001

Journal of Biological Chemistry

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were first found as a result of post-translational modifications of Arg residues, mainly in DNA-and RNA-binding proteins (1). Further studies established that methylated L-Arg molecules also occur freely in body fluids (2-5) and various tissues (6, 7). Although degradation of Arg-methylated proteins has been shown to be a source of free MMA and ADMA (8, 9), a direct methylation of L-Arg via a so far unknown pathway has also been suggested (10, 11). The y ϩ membrane transporter system is apparently involved in the uptake or release of MMA and ADMA (11,12).Nitric-oxide synthase (NOS), which generates the free radical NO from L-Arg, exists in at least three isoforms that are involved in different biological processes. The two constitutive isoforms, nNOS (neuronal) and eNOS (endothelial), produce NO for neurotransmission and cardiovascular regulation, respectively, and are predominantly regulated by cytosolic Ca 2ϩ levels (13). It is well established that MMA and ADMA are endogenous competitive inhibitors of NOS (9,11,14). The intracellular L-Arg levels and the corresponding K m value of NOS are such that the production of NO should not depend on L-Arg supplementation (15). However, in biological studies, the addition of L-Arg increased the production of NO (16), a phenomenon called the "Arg paradox." In this context, it may be noted that recent in vitro studies have shown that the concomitant presence of varying, yet physiologically relevant, levels of MMA and ADMA dramatically affects NOS activity (17).The enzyme dimethylargininase (L-N ,N -dimethylarginine dimethylaminohydrolase (DDAH), EC 3.5.3.18) hydrolyzes both MMA and ADMA to L-citrulline (L-Cit), CH 3 NH 2 , and (CH 3 ) 2 NH, respectively (18). It has been established that DDAH regulates NOS by controlling the levels of MMA and ADMA (10). DDAH exists in at least two isoforms (19), both of which are cytosolic proteins (20). Whereas DDAH-1 is expressed predominantly in tissues containing nNOS (21-25), DDAH-2 is mainly found in tissues expressing eNOS (19,25). The role of DDAH-1 in the regulation of nNOS is also evident from recent studies showing that both proteins are up-regulated in injured neurons (26).The increased levels of MMA and ADMA in plasma and urine have been found in several diseases that are linked to vascular dysfunction characterized by low NO levels, e.g. uremia, atherosclerosis, hypercholesterolemia, diabetes mellitus, hypertension, and homocysteinemia (27). Moreover, increased levels of ADMA in plasma have been discussed as a risk factor for atherosclerosis (3,4,9). In contrast, NO overproduction apparently leads to diseases such as septic shock and migraine. In these cases, clinical studies using MMA as a drug have been or are currently being conducted (28,29).The above studies clearly indicate the importance of DDAH in NO metabolism. Previously, we have demonstrated that DDAH-1 from bovine brain is a monomeric (31.2 kDa) Zn(II)-containing protein (30) and that the Zn(II) is not involved in the * This work was supported in part by the Olga...

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Section: Protein Purificationmentioning

confidence: 99%

Structural and Functional Characterization of the Zn(II) Site in Dimethylargininase-1 (DDAH-1) from Bovine Brain

Knipp¹,

Charnock²,

Garner³

et al. 2001

Journal of Biological Chemistry

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“…Polymers, sugar residues and even graphite (Pan et al, 2004) were attached to antibodies and proteins. A wide range of applications is known: clinical (Yavuz and Denizli, 2005;Acevedo et al, 2006), agricultural and food-chemical (Raviyan et al, 2003;Tumturk et al, 1999), affinity-and ion-exchange chromatography-related (Ruhn et al, 1994;Pessela et al, 2006;van Sommeren et al, 1993) and biotechnological (Kumada et al, 2006;Ong et al, 1991;Ong et al, 1989;Reetz et al, 1996).…”

Section: Immobilization On Beadsmentioning

confidence: 99%

Current Advances in Antibody Immobilization on Different Surfaces and Beads

Hahn¹,

Bakry²,

Huck³

et al. 2008

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Antibody immobilization is of considerable interest for miscellaneous fields of interest, e.g. detecting biomarkers in cancer diagnostics, e.g. prostate specific antigen (PSA) for prostate cancer and pathogens like Escherichia coli, a foodborne pathogen. For this purpose, specific antibody captures are required and all of them should guarantee highest reproducibility and capacity possible. Especially in clinical applications very complex and specific immunoassays are needed. However, the complexity of pinning antibodies necessitates particular methods concerning the immobilization technique itself-because of their structure and specificity-and of course the analyzing methods. Glass, polymers, gold and even fullerene beads have been used as carriers, treated with various spacers and activation steps within the last few years. Furthermore, considerable attention is drawn to the regeneration of matrices and their capabilities for further antibody attachment. Commonly applied verification tools for the detection of the trapped antibodies include impedance spectroscopy (IS), atomic fluorescence microscopy (AFM), electrophoresis (EP), enzyme linked immunosorbent assays (ELISA), quartz crystal microbalance (QCM) and infrared spectroscopy (IR). Matrix assisted laser desorption/ionization-time-offlight mass spectrometry (MALDI-TOF/MS) is also a useful tool for proving the success of new antibody fixing procedures. Immobilization techniques have been improved concerning their reproducibility, binding capacity and high specificity. In this review, recent developments of antibody (Ab) immobilization are summarized, the individual applications are mentioned, advantages and disadvantages are discussed in detail.

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“…Since the peptide library did not contain lysine, the coupling chemistry was selected to exploit a single primary amine which was incorporated on the C-terminus of the peptide. This lysine reacts with an N-Hydroxysuccinimide (NHS) activated Sepharose resin (van Sommeren et al, 1993), forming a peptide bond. The flexibility of chemical synthesis allowed directed coupling to proceed only through the C-terminal lysine residue, as the peptide has a blocked (acetylated) amino terminus.…”

Section: Base Matrix Selection and Immobilization Chemistrymentioning

confidence: 99%

Development and validation of an affinity chromatography step using a peptide ligand for cGMP production of factor VIII

Kelley¹,

Tannatt²,

Magnusson

et al. 2004

Biotech & Bioengineering

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An affinity chromatography step was developed for purification of recombinant B-Domain Deleted Factor VIII (BDDrFVIII) using a peptide ligand selected from a phage display library. The peptide library had variegated residues, contained both within a disulfide bond-constrained ring and flanking the ring. The peptide ligand binds to BDDrFVIII with a dissociation constant of f1 AM both in free solution and when immobilized on a chromatographic resin. The peptide is chemically synthesized and the affinity resin is produced by coupling the peptide to an agarose matrix preactivated with N-hydroxysuccinimide. Coupling conditions were optimized to give consistent and complete ligand incorporation and validated with a robustness study that tested various combinations of processing limits. The peptide affinity chromatographic operation employs conditions very similar to an immunoaffinity chromatography step currently in use for BDDrFVIII manufacture. The process step provides excellent recovery of BDDrFVIII from a complex feed stream and reduces host cell protein and DNA by 3 -4 logs. Process validation studies established resin reuse over 26 cycles without changes in product recovery or purity. A robustness study using a factorial design was performed and showed that the step was insensitive to small changes in process conditions that represent normal variation in commercial manufacturing. A scaled-down model of the process step was qualified and used for virus removal studies. A validation package addressing the safety of the leached peptide included leaching rate measurements under process conditions, testing of peptide levels in product pools, demonstration of robust removal downstream by spiking studies, end product testing, and toxicological profiling of the ligand. The peptide ligand affinity step was scaled up for cGMP production of BDDrFVIII for clinical trials. B 2004 Wiley Periodicals, Inc.

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Comparison of three activated agaroses for use in affinity chromatography: Effects on coupling performance and ligand leakage

Cited by 29 publications

References 16 publications

Structural and Functional Characterization of the Zn(II) Site in Dimethylargininase-1 (DDAH-1) from Bovine Brain

Structural and Functional Characterization of the Zn(II) Site in Dimethylargininase-1 (DDAH-1) from Bovine Brain

Current Advances in Antibody Immobilization on Different Surfaces and Beads

Development and validation of an affinity chromatography step using a peptide ligand for cGMP production of factor VIII

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