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Biomedical subjects

D F Young

Publications and source records attributed to D F Young.

65 records · Page 4Linked to original sources
PubMed1971

Hereditary bilateral acoustic neuroma (central neurofibromatosis).

Fifty-five members of a kindred have bilateral acoustic neuroma. The trait is autosomal dominant with high penetrance. Onset is about age 20; survival varies from two to 42 years. In screening members, vestibular abnormalities are most consistently observed, but complete audiometric studies and brain scan are important, because no single study is diagnostically sufficient in all cases. Peripheral signs suggesting neurofibromatosis are rare, but other CNS tumors, often asymptomatic, are common suggesting this trait may represent a "central" form of neurofibromatosis.

Adolescent↗
PubMedDate not supplied

Thiolate and phosphorothioate functionalized fluoresceins and their use as fluorescent labels.

We report the syntheses of two new fluorescein derivatives, 3',6'-dihydroxy-3-oxo-2-[(phosphonothio)-acetyl]spiro[isobenzof uran- 1(3H),9'-9H-xanthene]-6-carboxylic acid hydrazide, disodium salt, a phosphorothioate fluorescein, and 3',6'-dihydroxy-3-oxo-2-(mercaptoacetyl)spiro[isobenzofuran-1(3H), 9'-9H- xanthene]-6-carboxylic acid hydrazide, a mercaptoacetyl fluorescein. The latter is derived from the first compound by hydrolysis of the phosphate. Direct nonenzymatic labeling of the maleimide-derivatized IgG molecule by the novel mercaptoacetyl fluorescein is discussed. We also present a new method of bioconjugating phosphorothioate-functionalized fluorophores to a maleimide-derivatized protein, based on the alkaline phosphatase-catalyzed hydrolysis of the S-P bond of the phosphorothioate and the concomitant liberation of the fluorophore thiolate. This last species reacts in situ with the maleimide on the protein. A high degree of conjugation control is achieved in that modulation of the stoichiometry of the label and enzyme results in incorporation from seven to eight fluorophores per protein, depending on the ratio of the phosphorothioate fluorescein to alkaline phosphatase. The quantum yield of the mercaptoacetyl fluorescein relative to 6-carboxyfluorescein is 0.22 and lambda exc = 494 nm and lambda em = 517 nm.

Alkaline Phosphatase↗
PubMedDate not supplied

Thermally stabilized immunoconjugates: conjugation of antibodies to alkaline phosphatase stabilized with polymeric cross-linkers.

A method of conjugating poly(glutamic acid) poly(phosphorothioate)-cross-linked alkaline phosphatase to maleimide-derivatized immunoglobulin is described. Intramolecular autocatalyzed cross-linking of alkaline phosphatase at 2:1 to 4:1 polymer:enzyme ratios introduced 32-68 thiolates on the surface of the enzyme. Depending on the stoichiometry of polymer to enzyme, the cross-linked alkaline phosphatase retained 75-90% of its native catalytic activity. The cross-linked thiolate-functionalized alkaline phosphatase was conjugated to maleimide-derivatized immunoglobulin. Compared to a control prepared using non-cross-linked alkaline phosphatase, these conjugates were smaller in size and more stable to heat. The enzymatic activity of the cross-linked conjugates after incubation at 45 degrees C and pH 7.5 for 25 days was 35% higher than those of the highest-activity control conjugates. The conjugation process could be controlled by varying the stoichiometries of poly(glutamic acid) poly(phosphorothioate), alkaline phosphatase, and immunoglobulin.

Alkaline Phosphatase↗
PubMedDate not supplied

Alkaline phosphatase activatable polymeric cross-linkers and their use in the stabilization of proteins.

We report the synthesis of polymeric cross-linking agents, poly(glutamic acid) poly(phosphorothioates), and their use in the cross-linking and stabilization of proteins upon treatment with alkaline phosphatase. We have shown that poly(phosphorothioates) are excellent substrates of alkaline phosphatase, yielding thiolated polymers which react covalently with electrophilic groups introduced into the proteins. Three proteins of different structure and function were cross-linked using this method: calf intestinal alkaline phosphatase, glucose oxidase (Aspergillus niger), and (R)-phycoerythrin. The cross-linking of alkaline phosphatase is self-catalyzed since this enzyme catalyzes the hydrolysis of phosphates, unmasking thiolates which react with the maleimide prederivatized alkaline phosphatase. Incubation of buffered solutions of native alkaline phosphatase at 45 degreesC for 7-14 days resulted in a 35% higher loss of enzymatic activity compared to that of cross-linked enzyme. The effect of cross-linking glucose oxidase is even more notable, ranging from 800% stabilization at 37 degrees C and pH 9.0 to 3000% at 37 degrees C and pH 7.4. (R)-Phycoerythrin cross-linked with 1-3 equiv of poly(phosphorothioates) and incubated at 45 degrees C for 45 days was 20% more fluorescent than the native (R)-phycoerythrin subjected to the same conditions. The stabilizing effect of cross-linking was confirmed by comparing the rate of loss of quaternary structure of the cross-linked (R)-phycoerythrin with that of the native protein.

Alkaline Phosphatase↗