Site-specific modification of hemoglobin by methyl acetyl phosphate

Ueno, Hiroshi; Pospischil, M A; Manning, James M.; Kluger, Ronald

doi:10.1016/0003-9861(86)90648-x

Cited by 33 publications

(25 citation statements)

References 6 publications

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“…Indeed, our recent studies (12) and investigations by Manning and co-workers (Ref. 10 and reference therein) show that both agents acetylate ␤Lys-82 in the 2,3-DPG binding pocket (10,12). Based on this spectral comparison and in light of our recent study of asprin acetylation (12), we assign resonances 1, 3, and 6 to acetyl ␤Lys-59, acetyl ␣Lys-90, and acetyl ␤Lys-82, respectively.…”

Section: Resultsmentioning

confidence: 99%

“…Detection utilizing the relatively weak polarization transfer between the carbonyl 13 C and methyl protons of the acetyl group was facilitated by the high mobility of the acetyllysine side chains, and the carbonyl resonances were found to yield greater shift dispersion than the methyl resonances observed using [2-13 C]acetyl-labeled precursors. The 1 H- 13 C HMQC spectrum of COHbA acetylated using a 10-fold molar excess of [1][2][3][4][5][6][7][8][9][10][11][12][13] C]MAP is shown in Fig. 1A.…”

Section: Resultsmentioning

confidence: 99%

“…10 and 11 and references therein). No residues of the ␣-chain were reported modified in the same study (10). The modification of HbS by MAP was reported to yield adduct hemoglobin with oxygen affinity similar to that of hemoglobin A but with a reduced gelation tendency.…”

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confidence: 91%

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Acetylation of Human Hemoglobin by Methyl Acetylphosphate

Xu¹,

Labotka²,

London³

1999

Journal of Biological Chemistry

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Biophys. Acta 1432, 333-349). Blocking the 2,3-DPG binding site with inositol hexaphosphate (IHP) resulted in a selective reduction in intensity of adduct resonances, presumably corresponding to residues located in the 2,3-DPG binding cleft. The pattern of residue protection appeared to be identical to that observed in our previous study using IHP and labeled aspirin. Pre-acetylation of hemoglobin using unlabeled MAP, followed by acetylation with [1-13 C]aspirin indicated a general protective effect, with the greatest reduction of intensity for resonances corresponding to acetylated residues in the 2,3-DPG binding site. These studies indicated that both MAP and aspirin exhibit similar, although not identical, acetylation profiles and target primarily the ␤Lys-82 residue in the 2,3-DPG binding site, as well as sites such as ␤Lys-59 and ␣Lys-90, which are not located in the ␤-cleft of hemoglobin.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

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Acetylation of Human Hemoglobin by Methyl Acetylphosphate

Xu¹,

Labotka²,

London³

1999

Journal of Biological Chemistry

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“…The work in Manning's laboratory with Pospischil and Ueno established that methyl acetyle phosphate preferentially acetylates amino groups in the DPG-binding site of human hemoglobin, making it a potentially less toxic alternative to cyanate (16). The reagent is highly selective, reacting mainly with two of the 24 amino groups per alp dimer, the €-amino group of P-lys-82, and the a-amino group of the P-Nterminal valine (3,18). Further work from Manning and coworkers indicated that methyl acetyl phosphate is an effective anti-sickling agent and can also be used as a general acetylating agent for identifying sites that bind anions (19,20).…”

Section: Methyl Acetyl Phosphate and Hemoglobinmentioning

confidence: 99%

“…We have been developing methyl acyl phosphates as a new type of highly specific reagent for the modification of proteins (1)(2)(3)(4). We have found convenient routes for preparing a variety of acyl derivatives and have demonstrated the utility of these compounds in specifically cross-linking human hemoglobin so that the modified protein can be used in medical applications, such as carrying oxygen in a transfusion medium (5)(6)(7)(8).…”

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1994 Syntex Award Lecture: Anionic Electrophiles, Protein Modification, and Artificial Blood

Kluger¹

1994

Can. J. Chem.

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Monomethyl esters of acyl phosphates (mixed anhydrides of carboxylic and phosphoric acids) are being developed as site-directed acylating agents for amino groups in proteins. In an illustrative application, these anionic materials are shown to bind to a positively charged region of hemoglobin where they convert amino groups to amides. Bifunctional acyl methyl phosphates cross-link hemoglobin to give a variety of products, some of which have the oxygen-binding properties anticipated for materials that can serve as an alternative to red cells in transfusions. Higher yields of desired products result from the use of a trifunctional analogue. Kinetic patterns of the reactions of a series of alkyl amines and methyl aroyl phosphates indicate that the transition state for formation of the amide involves almost complete development of positive charge on nitrogen.

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Site-specific modification of hemoglobin by methyl acetyl phosphate

Cited by 33 publications

References 6 publications

Acetylation of Human Hemoglobin by Methyl Acetylphosphate

Acetylation of Human Hemoglobin by Methyl Acetylphosphate

1994 Syntex Award Lecture: Anionic Electrophiles, Protein Modification, and Artificial Blood

Covalent Inhibitors of the Gelation of Sickle Cell Hemoglobin and Their Effects on Function

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