The Isolation of a Pancreatic Insulinase

Lewis, U. J.; Thiele, Elizabeth H.

doi:10.1021/ja01560a075

Cited by 27 publications

(5 citation statements)

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“…There is also much evidence that it is a proteolytic enzyme probably related to chymotrypsin and that it has a great specificity for insulin. When this had been established, and when many other proteolytic enzymes from the pancreas had been eliminated on the basis of their specificity (trypsin and elastase) or their physical properties (carboxypeptidase, leucine aminopeptidase, and the 'insulinase' described by Lewis & Thiele, 1957), it became of importance to show that APP was distinct from chymotrypsin. The evidence that this is so is based partly on physical and chemical properties.…”

Section: Discussionmentioning

confidence: 99%

Interaction of pancreas extracts with bovine insulin. 1. Purification and chemical properties

Czerkawski¹,

Bingle²

1964

Biochemical Journal

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This paper is concerned with pancreatic substances that can antagonize insulin. Previous work (Marks & Young, 1939; Thorogood & Zinmmermann, 1945; Mirsky, Futterman, Wachman & Perisutti, 1951) has already suggested that insulin antagonists exist in the pancreas. Evidence has been produced for the identity of these antagonists with glucagon, but there is other work suggesting that the pancreas may produce substances that can antagonize insulin in such tissue as muscle (Young, 1962; Antoniades, Renold, Dagenais & Steinke, 1960) when glucagon is without effect (Randle, 1958). Antoniades et al. (1960) suggested that insulin might be combined with a basic protein in pancreas. The studies reported below are an attempt to isolate and characterize at least one of the bovine pancreatic substances that is capable of antagonizing insulin in a biological system such as rat diaphragm. MATERIALS AND METHODS Materials. DEAE-cellulose and CM-cellulose were obtained from Whatman (W. R. Balston Ltd.). Before use DEAE-cellulose was treated with 2N-NH3 and 2M-NaCl; CM-cellulose was treated with 2N-acetic acid and 2M-NaCl; both were washed well with distilled water. The bovine insulin (22-2 i.u./mg.) was obtained from Boots Pure Drug Co. Ltd., Nottingham. The proteolytic enzymes trypsin and oc-chymotrypsin were recrystallized preparations, obtained from L. Light and Co. Ltd., Colnbrook, Bucks. The pancreatic elastase was prepared by the method of Hall & Czerkawski (1959). Lilley & Valiance-Owen (1961) have shown that dialysis tubing contains substances capable of antagonizing insulin, and that these substances can be removed by boiling the tubing in two changes of distilled water. It was deemed wise to take the same precaution with the 1 in. tubing (Visking) used in the present work. Unless otherwise stated all the reagents used were of AnalaR quality. Electrophoresis. The purity of the fractions was frequently checked by use of electrophoresis at high voltage. on paper and at low voltage on cellulose acetate (Oxoid Division of Oxo Ltd., London, E.C. 4) as supporting medium. The electrophoresis was usually performed in a 75 mM-barbiturate (veronal) buffer solution, pH 8-6, which,

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Section: Discussionmentioning

confidence: 99%

Interaction of pancreas extracts with bovine insulin. 1. Purification and chemical properties

Czerkawski¹,

Bingle²

1964

Biochemical Journal

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“…Miyada & Tappel (1956) also reported the activity of a commercial proteolytic preparation of fungal origin, Rhozyme P-il. Lewis & Thiele (1957) showed by chromatography that their crystalline pancreatic elastase was heterogeneous and that, of the five components separated, only one component possessed elastase activity. Grant & Robbins (1957), using pancreatic elastase which had been partially purified by adsorption on to elastin, showed that it had strong proteolytic activity on proteins other than elastin.…”

mentioning

confidence: 99%

The chemistry of connective tissues. 5. The elastase activity of proteolytic enzymes

Thomas

Partridge

1960

Biochemical Journal

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“…Practically all the work of the last 10 years regarding elastase involves the pancreatic enzyme. Lewis' (254) crude crystalline elastase contains at least five components, only one of them elastolytic (255). Czerkawski and Hall (77) found yet a third component, a denatured form of the mucolytic factor which can suppress the synergistic effect and thus lower apparent activity.…”

Section: Historymentioning

confidence: 99%

“…Cellulose ion exchangers DEAE (255) and CM (328,425) eliminated certain activities, and Bagdy and Banga (lo), after elution of the adsorbed enzyme from a zeolite resin with alcoholic ammonium acetate and reprecipitation, obtained a product of greater purity and higher elastase activity than Banga's (20) crystalline enzyme. By adsorption on an Amberlite IRC-50 resin and subsequent elution with phosphate buffer, pH 6.0.…”

Section: Historymentioning

confidence: 99%

“…Slower splits in both the A and the B chain occurred at a Tide variety of pepttide bonds involving neutral amino acids with aliphatic side chains (328). A non-elastolytic insulinase has, however, been separated from a crude crystalline elastase preparation (255). Because of the apparent specificity for aromatic amino acids as well as other reasons (cf.…”

Section: Y Specijcity (A) Pancreatic Elastasementioning

confidence: 99%

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