Studies of a reactive histidine of diphosphopyridine nucleotide-linked isocitrate dehydrogenase from bovine heart

Fan, Chinan C.; Stegeman, William J.; Plaut, G.W.E.

doi:10.1016/0003-9861(77)90333-2

Cited by 10 publications

(2 citation statements)

References 23 publications

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“…The relationship becomes linear at high concentrations of dye where, [ANSI S [protein]. This be- haviour differs from the rectangular hyperbola observed with other proteins [21,27,281. A linear increase in fluorescence with increasing fluorophore is indicative of partitioning [29].…”

Section: Resultsmentioning

confidence: 85%

“…The quantum yield of the protein-bound ANS was determined by comparison with ANS in 100% ethanol as the reference (quantum yield c$ of 0.37 [21]). The areas under the emission spectra and the 360-nm absorbance of ANS were then used to calculate the quantum yield by the method of Parker and Rees [22] (1) where A is the area under the corrected emission spectrum, D is the absorbance at the exciting wavelength.…”

Section: Methodsmentioning

confidence: 99%

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Binding of 1‐Anilinonaphthalene‐8‐Sulfonic Acid to α‐crystallin

Stevens¹,

Augusteyn²

1997

European Journal of Biochemistry

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a-Crystallin was found to exhibit a time-dependent uptake of the hydrophobic probe, l-anilinonaphthalene-8-sulfonic acid (ANS), similar to that typically observed with lipid membranes. Analysis of the interaction of ANS with a-crystallin revealed two types of interactive processes, partitioning and binding. The predominant process involved partitioning, with a coefficient of 300 M-'. The binding component had the following characteristics: 1 binding site124 subunits and a Kd of about 9 pM. The binding was unaffected by the number of subunits used in the assembly of the a-aggregate, since both the an,-and a,-forms had similar binding characteristics. No discernible differences were observed in the binding of ANS to homopolymers of aA and aB subunits, suggesting that the hydrophobic sites to which ANS bound were similar in both the A and B subunits. The majority of the fluorescence was lost when the protein was incubated in 3 M urea, a concentration of denaturant where the protein is still intact, suggesting that the ANS binding sites are located near the surface of the protein. The decrease was attributed to a decrease in the quantum yield of the bound dye.Keywords: a-crystallin; I-anilinonaphthalene-8-sulfonic acid; quaternary structure; hydrophobic interaction.a-Crystallin constitutes up to 50% of the total proteins which comprise the bulk of the mammalian eye lens. Its relatively constant proportion in most species suggests that it has an important role in the lens. The absence of any tissue turnover necessitates that it remains functional for the lifetime of the organism, in the case of humans in excess of 70 years. In cases where the protein is modified or structurally disrupted impairment of vision results, usually in the form of a cataract. Recently, it has been shown that a-crystallin is a member of the small heat-shockprotein family and may function as a chaperone [I]. It has been suggested that one of its major functions is to prevent age-related aggregation of lens proteins, thereby maintaining lens transparency [2]. The mechanisms for this function have not been delineated but they probably involve the surface of a-crystallin which has been shown to be very hydrophobic [3, 41. Knowledge of the location of these hydrophobic sites would be of enormous benefit in determining how a-crystallin functions as a molecular chaperone, as well as providing insight into its structure.a-Crystallin is a polymeric protein which consists of two types of subunits, A and B, both of which have a mass around 20 kDa [5]. Their amino acid sequences are closely related and have changed very slowly during the course of evolution, reflecting a structural constraint on the protein. This also suggests that it has an important role in maintaining interactions necessary for the formation of a transparent lens. Both polypeptides are subject to various chemical and enzymatic modifications such as oxidation, phosphorylation, deamidation, glycation, and C-terminal degradation 161. These modifications are thought to

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Section: Resultsmentioning

confidence: 85%

Section: Methodsmentioning

confidence: 99%

Binding of 1‐Anilinonaphthalene‐8‐Sulfonic Acid to α‐crystallin

Stevens¹,

Augusteyn²

1997

European Journal of Biochemistry

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Histidin im aktiven Zentrum von Enzymen

Schneider¹

1978

Angew. Chem.

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Histidine in Enzyme Active Centers

Schneider

1978

Angew. Chem. Int. Ed. Engl.

119

100

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The presence of histidine in the active center of an enzyme can be demonstrated by kinetic measurements, chemical modification, NMR spectroscopy or X-ray structure analysis. Histidine is the only naturally occurring amino acid to contain an imidazole residue as a side chain. The role of histidine in enzyme catalysis depends, inter alia, upon the special features of the imidazole residue: it thus tends to form hydrogen bonds, combines donor and acceptor properties and can take part in either nucleophilic or base catalysis. In some of these enzymes the position of each atom is known; however, the theories as to how the catalysis proceeds at a molecular level are controversial.

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Studies of a reactive histidine of diphosphopyridine nucleotide-linked isocitrate dehydrogenase from bovine heart

Cited by 10 publications

References 23 publications

Binding of 1‐Anilinonaphthalene‐8‐Sulfonic Acid to α‐crystallin

Binding of 1‐Anilinonaphthalene‐8‐Sulfonic Acid to α‐crystallin

Histidin im aktiven Zentrum von Enzymen

Histidine in Enzyme Active Centers

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