Unfolding kinetics of bovine trypsinogen.


Abstract

The unfolding kinetics of bovine trypsinogen were studied by a fluorescence-detected stopped-flow technique at pH 5.8. Trypsinogen unfolding appeared to be a rather complex reaction. Two phases, fast (with a time constant in the millisecond range) and slow, were detected in the range 2-7 M guanidium chloride (GdmCl). The natural logarithm of the rate constant of the slow phase exhibited strong dependence on [GdmCl], changing from hundreds of seconds at low denaturant concentration to about 20 ms at 7 M GdmCl. The curvature of this dependence further suggests a complex mechanism of unfolding. Generally, similar kinetics were observed for the trypsinogen.Ca complex. Small differences could be noticed, however, for the fast phase. In agreement, Ca2+ influenced only this stage of the reaction. Analysis of the dependence of the time constant of the fast phase on [CaCl2] indicates that at 4 M GdmCl, trypsinogen.Ca unfolds about sixfold slower than free zymogen, and that native trypsinogen at 4 M GdmCl still exhibits high affinity for Ca2+. Limited data on trypsin unfolding show virtually an identical dependence of the slow phase on [GdmCl]; the fast phase, however was not observed. Moreover, in the 3-4.5 M GdmCl range, a separate phase was detected. It is postulated that this phase is a manifestation of the activation-domain unfolding. The Eyring plots for the fast phase of . trypsinogen and trypsinogen.Ca unfolding are linear, indicating little change in heat capacity for this stage of reaction. The slow step of unfolding, however, shows significant curvature which indicates a substantial increase in heat capacity. Study holds ProTherm entries: 9882 Extra Details: trypsinogen; unfolding kinetics; transition state;,denaturation; guanidinium chloride

Submission Details

ID: sBnWrZ8r

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:39 p.m.

Version: 1

Publication Details
Otlewski J;Sywula A;Kolasinski M;Krowarsch D,Eur. J. Biochem. (1996) Unfolding kinetics of bovine trypsinogen. PMID:9022687
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P
 

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)


Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Cationic trypsin P00760 TRY1_BOVIN