The effect of bile salts on the esterolytic assay of trypsin

Calvert, G. D.; Legg, E F

doi:10.1016/0009-8981(77)90148-6

Cited by 3 publications

(3 citation statements)

References 13 publications

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“…Bile acids play an important role in intestinal absorption of lipids 30 , but also may promote protein denaturation and potentiate the activity of digestive proteases 31 , 32 . Post-prandial free bile acid concentrations in the human small intestine reach up to 10 mM 6 .…”

Section: Resultsmentioning

confidence: 99%

Gastrobodies are engineered antibody mimetics resilient to pepsin and hydrochloric acid

2021

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Protein-based targeting reagents, such as antibodies and non-antibody scaffold proteins, are rapidly inactivated in the upper gastrointestinal (GI) tract. Hydrochloric acid in gastric juice denatures proteins and activates pepsin, concentrations of which reach 1 mg/mL in the mammalian stomach. Two stable scaffold proteins (nanobody and nanofitin), previously developed to be protease-resistant, were completely digested in less than 10 min at 100-fold lower concentration of pepsin than found in the stomach. Here we present gastrobodies, a protein scaffold derived from Kunitz soybean trypsin inhibitor (SBTI). SBTI is highly resistant to the challenges of the upper GI tract, including digestive proteases, pH 2 and bile acids. Computational prediction of SBTI’s evolvability identified two nearby loops for randomization, to create a potential recognition surface which was experimentally validated by alanine scanning. We established display of SBTI on full-length pIII of M13 phage. Phage selection of gastrobody libraries against the glucosyltransferase domain of Clostridium difficile toxin B (GTD) identified hits with nanomolar affinity and enzyme inhibitory activity. Anti-GTD binders retained high stability to acid, digestive proteases and heat. Gastrobodies show resilience to exceptionally harsh conditions, which should provide a foundation for targeting and modulating function within the GI tract.

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Section: Resultsmentioning

confidence: 99%

Gastrobodies are engineered antibody mimetics resilient to pepsin and hydrochloric acid

2021

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“…It has been observed in earlier studies that related bile salts affect the esterolytic activity of thrombin (EC 3.4.21.5) (Curragh & Elmore, 1964;Exner & Koppel, 1973) and trypsin (Calvert & Legg, 1977).…”

Section: Effect Ofaggregationmentioning

confidence: 87%

“…For thrombin such effects have been proposed to .result from both conformational changes occurring in the enzyme or enzyme-substrate complex and the _0 _4binding of solvent molecules to the enzyme (Curragh & Elmore, 1964). For trypsin it has been suggested that the bile salts affect the ester substrate rather than the enzyme itself (Calvert & Legg, 1977 (Chao, 1978).…”

Section: Effect Ofaggregationmentioning

confidence: 99%

Studies on the temperature-dependent autoinhibition of human plasma kallikrein I

Levison¹,

Tomalin²

1982

Biochemical Journal

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At 37 degrees C, human plasma kallikrein I follows Michaelis-Menten behaviour and exhibits a normal linear relationship between the initial velocity of hydrolysis of Ac-Pro-Phe-Arg-OMe,HCl and enzyme concentration in the range 0--150 pM. At temperatures of 30 degrees C and below substantial deviations from linearity are observed over the same enzyme concentration range. The temperature-dependent autoinhibition of kallikrein I activity is reversible and is not due to low-molecular-weight endogenous inhibitors or cofactors. The kinetic effect is apparently due to aggregation and can be abolished by the addition of sodium deoxycholate.

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Importance of bile in regulation of intraluminal proteolytic enzyme activities in the rat

Green

Nasset

1980

Gastroenterology

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The effect of bile salts on the esterolytic assay of trypsin

Cited by 3 publications

References 13 publications

Gastrobodies are engineered antibody mimetics resilient to pepsin and hydrochloric acid

Gastrobodies are engineered antibody mimetics resilient to pepsin and hydrochloric acid

Studies on the temperature-dependent autoinhibition of human plasma kallikrein I

Importance of bile in regulation of intraluminal proteolytic enzyme activities in the rat

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