A-Chymotrypsinogen

Fig. 4. HPHIC of standard proteins on the weak hydrophobic columns. The SynChro-pack PROPYL column was 25x0.41 cm Poly (alkyl aspartamid)-silicas were packed into 20 x 0.46 cm columns. Sample 25 pi containing 25 pg of each protein in buffer A. Buffer A 1.8 mol/1 ammonium sulphate + 0.1 mol/1 potassium phosphate, pH 7.0. Buffer B 0.1 mol/1 potassium phosphate, pH 7.0. Gradient 40-min linear 0-100% buffer B. Flow rate 1 ml/min. Detection A220 = 1-28 a.u.f.s. Peaks a = cytochrome C, b = ribonu-clease A, c = myoglobin, d = conalbumin, e = neochymotrypsin, / = a-chymotrypsin, g - a-chymotrypsinogen A [48]...

Fractionated ammonium sulfate precipitaion of a-chymotrypsinogen (further fractions contain deoxyribonuclease, chymotrypsinogen B, ribonuclease, trypsinogen). 

Figure 4.8 Cation-exchange liquid chromatography of basic proteins. Column, Asahipak ES502C eluent, 20 min linear gradient of sodium chloride from 0 to 500 mM in 50 mM sodium phosphate buffer pH 7.0 flow rate, 1 ml min-1 temperature, 30 °C detection, UV 280 nm. Peaks 1, myoglobin from horse skeletal muscle (Mr 17 500, pi 6.8-7.3) 2, ribonuclease from bovine pancreas (Mr 13 700, pi 9.5-9.6) 3, a-chymotrypsinogen A from bovine pancreas (Mr 257 000, pi 9.5) and 4, lysozyme from egg white (Mr 14 300, pi 11.0-11.4). (Reproduced by permission from Asahikasei data)...

Fig. Z14. The activation of chymotrypsin via proteolytic cleavage, a) Chymotrypsinogen is transformed into the active forms of chymotrypsin n and a by trypsin and autoproteolysis, b) The N-terminal isoleucine residue Ile6 is particularly important for the activity of chymotrypsin. The positively charge NH2 group of llel6 interacts electrostatically with Aspl94 and stabilizes an active conformation of the catalytic center. After Stryer Biochemistry , with permission.

Figure 3-10 Estimation of the molecular mass of the polypeptide chain of the nitrogenase Fe-protein using SDS-poly-acrylamide electrophoresis from a set of four standard curves. The marker proteins are (1) catalase, (2) fumarase, (3) aldolase, (4) glyceraldehyde-phosphate dehydrogenase, (5) a-chymotrypsinogen A, and (6) myoglobin, (o) indicates position of azoferredoxin. From Nakos and Mortenson.195...

Figure 2.9 Hydrophobic-interaction chromatography of proteins. (A) Ammonium sulfate gradient from 2.16 to 0 M (B) ammonium sulfate and tetrabutylammonium bromide gradients from 2.16 to 0 M and from 0 to 10 mM, respectively (C) ammonium sulfate and tetrabutylammonium bromide gradients from 2.16 to 0 M and from 0 to 20 mM, respectively (D) ammonium sulfate and tetrabutylammonium bromide gradients from 2.16 to 0 M and from 0 to 40 mM, respectively. Chromatography conditions column, silica-bound polyether, 10 cm x 4.6 mm I.D. temperature, 25°C flow rate, 1 ml/min gradient, linear for 30 min background buffer, 50 mM phosphate, pH 6.5. Peaks a, cytochrome c b, ribonuclease A c, /3-lactoglobulin A d, lysozyme e, ovalbumin f, a-chymotrypsinogen A g, fetuin. (Reprinted from Ref. 45 with permission.)...

Fig. 6.18. Electrochromatogram of four basic proteins obtained by isocratic separation using a modified polychloromethylstyrene-based PLOT column (Reprinted with permission from [50]. Copyright 1999 Elsevier). Column 47 cm (active length 40 cm) x 20 pm, inner polymer layer 2 pm mobile phase 20% acetonitrile in 20 mmol/1 phosphate buffer pH 2.5 voltage -30 kV EOF velocity measured with dimethylsulfoxide (DMSO) -3.46 x 10"8 m2V ls 1, migration time for DMSO 3.10 min. Peaks a-chymotrypsinogen (1), ribonuclease (2), lysozyme (3), cytochrome C (4).

Activation of a-chymotrypsinogen by dissolution in 0,005 normal HC1, standardization to 0,1 molar CaCI2 and 0,1 molar borate buffer pH 8.0 separation of inactive precipitate after 24 h precipitation of Ca2t as sulfate. 

Protein-Pak SP-8HR (strong cation exchange, 8 /t particles) Protamine sulfate a-chymotrypsinogen A, cytochrome c and lysozyme 23... 

Waters SP-8HR (strong cation exchange, 8 fi particles) Low molecular weight dendrimers (PETMA4, 12, 36) a-chymotrypsinogen A and cyctochrome c 44... 

Battistel, E., Attanasio, F., Rialdi, G. (2000) Thermal stability of immobilized a-chymotrypsinogen, Journal of Thermal Analysis and Calorimetry 61(2), 513-525. 

Carbonic anhydrase, a-lactalbumin, trypsin inhibitor, oval-bumn, conalbumin, hemoglobin variants Ribonuclease, insulin, a-lactalbumin Lysozyme, a-chymotrypsinogen, ribonuclease A, cytochrome c... 

Denaturation horse heart cytochrome Denaturation whale myoglobin Denaturation a-chymotrypsinogen Precipitation sheep liver nucleoprotein at 40°C in 30 minutes... 

Titration curves for bovine a-chymotrypsinogen have been determined under a variety of conditions by Wilcox (1961). The results are summarized in Table IX. The spectrophotometric titration of the phenolic groups is shown in Fig. 4. 

Fig. 24. The average lifetime of a-chymotrypsinogen A as a function of water content (A). Left and right ordinate scales are for different methods of averaging the two components detected in analysis of the decay curves. From Fucaloro and Forster (1985).

Fig. 1. Chromatography of bovine pancreatic juice on DEAE-cellulose (anionic proteins) and Amberlite IRC-50 (cationic proteins) (1). RNAase, ribonuclease ChTg-a, chymotrypsinogen A Tg, trypsinogen ProCp-B and Cp-B, procarboxypeptidase B and carboxypeptidase B DNAase, deoxyribonuclease ProCp-A, procarboxypeptidase A.

EDTA) at room temperature were reduced using 4.8 mM dithiol. Under these reaction conditions a maximum of 0.75 disulfide residue per a-Chymotrypsinogen A molecule was reduced (2). The analysis for reduction of a-Chymotrypsinogen A was similar to that for trypsinogen. 

Rate constants k) are apparent rate constants based on total dithiol concentration. The calculations of rate constants are described in Methods section. The rate constants for trypsinogen and a-chymotrypsinogen A are from reference 2. 

The disulfide bond in a-chymotrypsinogen A is reduced about 2.3-fold faster using BMS and DMH than by DTT (Table I). A maximum of 0.75 disulfide group per a-chymotrypsinogen A molecule was reduced under the reduction conditions. The apparent rate constant for the reduction of disulfide bond in... 

BSA monomer, ovalbumin (chicken), P-lactoglobulin (bovine milk), serum albumin (human), carbonic anhydrase (bovine), L-glutamic dehydrogenase (bovine liver), a-chymotrypsin (bovine), a-chymotrypsinogen A (bovine), immunoglobulin (bovine milk), pepsin, trypsin (bovine), and heparin were from Sigma. RNase and lysozyme (egg white) were from Calbiochem. The recombinant human basic fibroblast... 

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