POTENTIAL ANTICANCER AGENTS.<sup>1</sup> LXVI. NON-CLASSICAL ANTIMETABOLITES. III.<sup>2</sup> 4-(IODOACETAMIDO)-SALICYLIC ACID, AN EXO-ALKYLATING IRREVERSIBLE INHIBITOR OF GLUTAMIC DEHYDROGENASE

Baker, B. R.; Lee, William W.; Tong, Ethel; Ross, Leonard O.

doi:10.1021/ja01478a034

Cited by 71 publications

(21 citation statements)

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“…At 10 mM BME, the interchain disulfide is half reduced, whereas the Y-2-Fab disulfide is only approximately 20% reduced. 2 It is also notable that a significant portion of the 25kD disulfide-linked Y-2-heavy chain is produced at higher BME concentrations, confirming that the interchain disulfide is reduced while the Y-2 disulfide remains intact. Since the Y-2-Fab disulfide bond is more stable than the interchain disulfide, Y-2-Fab should remain intact under physiological conditions.…”

Section: Quantitation Of Covalent Bondmentioning

confidence: 83%

“…This solution was allowed to incubate at 37 °C for periods up to 160 min. 2 In each case the half-lives for covalent attachment were a few minutes at 37 °C, pH 7.4, and the attached ligands migrated with the protein during denaturing (non-reducing) gel electrophoresis. The results for ligands 1 and 2, shown in Figure 2, are similar to the previous results for electrophiles 3 and 4 reacting with the free sulfhydryl form of this antibody (10).…”

Section: Determination Of Covalent Bond Formation Between the Cysteinmentioning

confidence: 99%

“…To determine the percentage of chelate covalently bound to the protein, reversibly bound chelate was separated from the protein on a denaturing (nonreducing) polyacrylamide gel. 2 The results for the four samples, reduced or non-reduced 2D12. Having formed the disulfide, we sought an indication of how stable the bond is.…”

Section: Quantitation Of Covalent Bondmentioning

confidence: 99%

“…After DTT reduction, we used amino-acid analysis to confirm the release of cysteine monomer from the Fab. 2 As test affinity labels we synthesized nucleophilic compounds 1 and 2 to compare with previously reported electrophiles 3 and 4 (Chart 1). The method of Weiss (25) was used to prepare 1 from thiocyanate, bromoacetate, and aminobenzyl-DOTA; Traut's reagent was used to prepare 2.…”

Section: Introductionmentioning

confidence: 99%

“…The method of Weiss (25) was used to prepare 1 from thiocyanate, bromoacetate, and aminobenzyl-DOTA; Traut's reagent was used to prepare 2. 2 …”

Section: Introductionmentioning

confidence: 99%

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Cysteinylated Protein as Reactive Disulfide: An Alternative Route to Affinity Labeling

et al. 2007

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Engineering the permanent formation of a receptor-ligand complex has a number of promising applications in chemistry, biology, and medicine. Antibodies and other proteins can be excellent receptors for synthetic ligands such as probes or drugs. Because proteins possess an array of nucleophilic sites, the placement of an electrophile on the synthetic ligand to react with a nucleophile on the macromolecule is a standard practice. Previously, we have used the site-directed incorporation of cysteine nucleophiles at the periphery of an antibody's binding site, paired with the chemical design of weakly electrophilic ligands, to produce receptor-ligand pairs that conjugate specifically and permanently ) Bioconjugate Chem.15, 1392-1402 and references therein). After protein expression in Drosophila S2 cells we found, as is frequently observed, that the engineered cysteine was reversibly blocked by disulfide linkage to a cysteine monomer (cysteinylated). Removal of the cysteine monomer requires some care because of the need to preserve other disulfide linkages in the protein. Here we report that cysteinylation can be used to advantage by treating the cysteine monomer as a leaving group and the protein disulfide as an electrophile with special affinity for thiols. Two ligands bearing thiol side chains were synthesized and incubated with the cysteinylated antibody Fab fragment 2D12.5 G54C, with the finding that both ligands become covalently attached within a few minutes under physiological conditions. The attachment is robust even in the presence of excess thiol reagents. This rapid, specific conjugation is particularly interesting for biomedical applications.

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Section: Quantitation Of Covalent Bondmentioning

confidence: 83%

Section: Determination Of Covalent Bond Formation Between the Cysteinmentioning

confidence: 99%

Section: Quantitation Of Covalent Bondmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

“…The method of Weiss (25) was used to prepare 1 from thiocyanate, bromoacetate, and aminobenzyl-DOTA; Traut's reagent was used to prepare 2. 2 …”

Section: Introductionmentioning

confidence: 99%

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Cysteinylated Protein as Reactive Disulfide: An Alternative Route to Affinity Labeling

et al. 2007

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Affinity Labelling of Protein Active Sites

1970

Ciba Foundation Symposium ‐ Molecular Properties of Drug Receptors

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Dynamic Covalent Capture: A Sensitive Tool for Detecting Molecular Interactions

Prins

2010

Ideas in Chemistry and Molecular Sciences

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POTENTIAL ANTICANCER AGENTS.¹ LXVI. NON-CLASSICAL ANTIMETABOLITES. III.² 4-(IODOACETAMIDO)-SALICYLIC ACID, AN EXO-ALKYLATING IRREVERSIBLE INHIBITOR OF GLUTAMIC DEHYDROGENASE

Cited by 71 publications

References 1 publication

Cysteinylated Protein as Reactive Disulfide: An Alternative Route to Affinity Labeling

Cysteinylated Protein as Reactive Disulfide: An Alternative Route to Affinity Labeling

Affinity Labelling of Protein Active Sites

Dynamic Covalent Capture: A Sensitive Tool for Detecting Molecular Interactions

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POTENTIAL ANTICANCER AGENTS.1 LXVI. NON-CLASSICAL ANTIMETABOLITES. III.2 4-(IODOACETAMIDO)-SALICYLIC ACID, AN EXO-ALKYLATING IRREVERSIBLE INHIBITOR OF GLUTAMIC DEHYDROGENASE

Cited by 71 publications

References 1 publication

Cysteinylated Protein as Reactive Disulfide: An Alternative Route to Affinity Labeling

Cysteinylated Protein as Reactive Disulfide: An Alternative Route to Affinity Labeling

Affinity Labelling of Protein Active Sites

Dynamic Covalent Capture: A Sensitive Tool for Detecting Molecular Interactions

Contact Info

Product

Resources

About

POTENTIAL ANTICANCER AGENTS.¹ LXVI. NON-CLASSICAL ANTIMETABOLITES. III.² 4-(IODOACETAMIDO)-SALICYLIC ACID, AN EXO-ALKYLATING IRREVERSIBLE INHIBITOR OF GLUTAMIC DEHYDROGENASE