Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.1 (chymotrypsin)
 10,938 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Steady-state kinetic measurements have shown that 8-azido-1,N6-ethenoadenosine 5'-triphosphate (8-N3-epsilon ATP) can be noncovalently bound to rabbit muscle fructose 1,6-bisphosphate aldolase with Ki = 0.075 mM at pH 8.5. This binding is purely competitive with substrate and occurs at the strong binding site for mononucleotides. Photoaffinity labeling of aldolase in the presence of 8-azido-1,N6-ethenoadenosine 5'-triphosphate results in inactivation of the enzyme. Aldolase is protected against modification in the presence of the inhibitors hexitol 1,6-bisphosphate or ATP. The labeling is saturable, and a good correlation is observed between the loss of enzymatic activity and the incorporation of 8-N3-epsilon ATP into aldolase. In addition, aldolase loses its ability to bind to phosphocellulose following modification. Digestion of labeled protein with trypsin, chymotrypsin, and cyanogen bromide revealed substantial modification of peptide 259-269. Thr-265 was identified as the residue that was covalently modified by 8-N3-epsilon ATP. On the basis of these results and other data we propose a model for the mononucleotide binding site.
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PMID:Photoaffinity labeling of rabbit muscle fructose-1,6-bisphosphate aldolase with 8-azido-1,N6-ethenoadenosine 5'-triphosphate. 365 92

The sorbitol permease enables the efflux of sorbitol from cultured rabbit papillary cells (PAP-HT25) in response to a reduction in osmolality. The anion transport inhibitor 5-nitro-2-(3-phenylpropylamino)benzoate (100 microM) inhibited efflux by 92%, and 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (0.5 mM) reduced efflux by 40%. 2,4-Dinitrobenzenesulfonate and p-chloromercuriphenylsulfonic acid had no effect. The protease trypsin (0.05 mg/ml) reduced sorbitol efflux by 53%, pronase (0.01 mg/ml) by 47%, and papain (0.1 mg/ml) by 49%; chymotrypsin had no effect. Sugars and sugar alcohols at different concentrations (10-200 mM) in the bathing solution did not influence sorbitol efflux. Determination of the osmotically induced influx of sugar alcohols showed that xylitol uptake was faster than that of sorbitol; 6-deoxysorbitol was slower; L-sorbitol, arabitol, galactitol, and 2-deoxysorbitol entered at the same rate as sorbitol; and maltitol did not enter the cells. Sorbitol and 6-deoxysorbitol at 9 mM competitively inhibited [14C]sorbitol influx by 24 and 32%, respectively, whereas xylitol, taurine, betaine, and myo-inositol showed no inhibition. We conclude that 1) a specific inhibitor of the permease was not found, 2) the sorbitol permease or associated regulator is a protein, and 3) the C-6 atom of sorbitol is important in the selectivity of the permease.
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PMID:Further characterization of the sorbitol permease in PAP-HT25 cells. 807 86

Differential scanning calorimetry was the method to investigate the thermostability of chymotrypsin. The transition temperature decreased by approx. 30 degrees C when the dry enzyme became highly hydrated. High degree of hydration corresponded to extensive conformational changes during protein denaturation, reflected by large enthalpy values. Sorbitol, lyophilized together with the enzyme, caused the destabilization of the complex within the whole range of water activities. When the enzyme was equilibrated through the apolar solvent, isooctane, stabilization of chymotrypsin was observed at high water activities, compared to equilibration in air. The presence of isooctane resulted in a remarkable stabilization of the chymotrypsin-sorbitol complex. A sorbitol concentration of 5 mmol/g of protein was prerequisite to induce stabilization when equilibrated through isooctane at high water activities. The transition enthalpy increased with increasing amounts of sorbitol. Different hydration isotherms were obtained for the air-equilibrated and solvent-equilibrated enzyme preparations. Increasing amounts of buffer salts within the chymotrypsin preparation caused the enhancement of both the temperature and the enthalpy of the transition at a water activity 0.97. Variations on the hydration of the preparations both offered the explanation to the thermal stability results and supported the evidence obtained from enzyme activity studies. Generally, the catalyst whose hydration was suppressed due to its environment exhibited low enzymatic activity.
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PMID:Calorimetric studies on solid alpha-chymotrypsin preparations in air and in organic solvents. 867 68

The effect of the addition of sorbitol on the activity and stability of enzymes was examined by monitoring transesterification reactions performed in organic media at various water activities (a(w) = 0.08 to 0.97). Lipases from Chromobacterium viscosum and Candida rugosa immobilized on celite, and chymotrypsin, free or immobilized on celite, were used. When the sorbitol-containing enzymes were employed, higher reaction rates and less hydrolysis were observed. Immobilization of chymotrypsin resulted in high activity and operational stability, while the nonimmobilized enzyme was stable only in the presence of sorbitol. The activity of all preparations diminished after washing them with pyridine to remove sorbitol. Furthermore, severe stability problems occurred in the preparations lacking sorbitol. Sorbitol treatment, even after removal of the sorbitol itself, improved the activity of nonimmobilized chymotrypsin relative to the washed control. On the other hand, washing to remove sorbitol had a negative effect on the activity of both coimmobilized lipase and coimmobilized chymotrypsin. Addition of a substrate analogue, N-acetyl-L-phenylalanine, to chymotrypsin yielded a preparation that exhibited higher activity than both the control and its sorbitol-containing counterpart. Differential scanning calorimetry measurements revealed that the chymotrypsin-sorbitol complex was stable against thermal denaturation, undergoing transition at a high temperature (89 degrees C). The transition temperatures of the substrate-containing chymotrypsin and of the control were identical (72 degrees C).
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PMID:Effects of sorbitol addition on the action of free and immobilized hydrolytic enzymes in organic media. 1862 33