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Thesis (Ph. D.)--University of Wisconsin--Madison, 1957. / Typescript. Abstracted in Dissertation abstracts, v. 17 (1957) no. 6, p. 1203-1204. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

The muscle aldolase from Ambystoma tigrinum has been purified 73-fold to a final specific activity of 13.2 units per mg. The purified enzyme appeared to be homogenous by ultracentrifugation and electrophoretic criteria. A molecular weight of 159,000 + 1000 was determined by gel filtration on Sephadex G-200 and high speed sedimentation equilibrium ultracentrifugation. The enzyme migrated identically with rabbit muscle aldolase when subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis and is apparently a tetramer of nearly identical subunits of approximately 40,000 MW. The catalytic constants of the salamander enzyme were similar to those reported for other muscle aldolases with the exception of the unusually low Fru-P2/FlP ratio.

Aldolase from human heart has been purified 128-fold to a final specific activity of 11.52 units per mg. The purification procedure employed column chromatography on phosphocellulose.

No description available.

A procedure is described for the purification of human erythrocyte aldolase (EC 4.1.2.13). The process involves a specific substrate elution of the enzyme from phosphocellulose followed by a reverse ammonium sulfate fractionation.

The fructose 1,6-bisphosphate type I aldolase from the thermophilic organism Thermoproteus tenax has been expressed and purified by heat treatment. The aldolase aldehyde substrate scope was investigated using electrospray mass spectrometry to detect the formation of any aldol products. Parameters affecting aldolase activity, including temperature, buffer pH and solvent additive were investigated. The synthesis of an aldehyde with an attached fluorescent reporter group was performed for potential use in the screening of mutant aldolases for aldol or Baylis-Hillman activity. The synthesis of 1-hydroxy-3-buten-2-one phosphate, an analogue of dihydroxyacetone phosphate, capable of participating in Baylis-Hillman reactions, was achieved in 5 steps from 3-buten-1-ol. This analogue was used in the investigation of the wild type aldolase and several mutant aldolases for Baylis-Hillman activity. X-ray crystallographic data was obtained for the wild type enzyme and the Trp144Leu mutant aldolase with 1-hydroxy-3-buten-2-one phosphate bound at their active sites. In the wild type aldolase, the substrate was found to bind in a similar manner to dihydroxyacetone phosphate, with the formation of a Schiff base with the Lys177 amino acid residue at the enzyme active site. In the Trp144Leu mutant aldolase, Lys177 has added in Michael fashion to the enone functionality of the bound substrate forming an enolate instead of forming a Schiff base. Both forms of the bound substrate are potentially capable of participating in Baylis-Hillman reactions. The enzymes have yet to be fully investigated for Baylis-Hillman activity.

Procedures for the isolation and characterization of Ascaris glyceraldehyde-3-phosphate dehydrogenase and actin are described. The properties of these proteins, including molecular weights, isoelectric points, kinetics, peptide maps, and amino acid compositions, are strikingly similar to the respective proteins from rabbit muscle.

No description available.

Mutationen im Gen der Aldolase B auf Chromosom 9 führen zur „hereditären“ Fructoseintoleranz (HFI), die nach einer längeren Fructoseexposition zu schweren Leber- und Nierenschäden bis hin zum völligen Versagen dieser Organe führen kann. Zum Ausschluß bzw. zur Bestätigung einer HFI werden seit ca. 20 Jahren hauptsächlich molekularbiologische Verfahren eingesetzt. Dabei zeigte es sich, dass wenige, regional unterschiedlich häufig vorkommende Mutationen (in Deutschland A149P und A174D) bei dem Großteil der Patienten für diese Erkrankung verantwortlich sind. Viele Laboratorien beschränken sich daher auf den Nachweis bzw. den Ausschluß dieser Mutationen. Ziel der vorliegenden Arbeit war es zu zeigen, dass auch bei den Patienten mit der Verdachtsdiagnose „hereditäre Fruktoseintoleranz“, bei denen die häufig vorkommen-den Mutationen A149P und A174D nur auf einem oder auf keinem der zwei Allele nachweisbar waren, die Diagnose mit molekularbiologischen Methoden durch den Nachweis seltener oder bisher nicht beschriebener Mutationen gesichert werden kann. Untersucht wurden 8 Patienten aus 5 Familien mit dieser Verdachtsdiagnose. Dabei handelte sich um Patienten mit klinischen Symptomen und in einem Fall enzymatisch gesicherter Diagnose. In allen Fällen mit Ausnahme von Patient III3 (Genotyp A174T/Wildtyp) konnten zwei Mutationen im Aldolase B-Gen nachgewiesen werden. Von den insgesamt identifizierten 8 Mutationen waren 3 zum Zeitpunkt der Durchführung der experimentellen Arbeiten in der Fachliteratur noch nicht beschrieben und erhöhen die Zahl der bekannten Mutationen im Aldolase B – Gen auf insgesamt 45.