The conformational stability and flexibility of insulin with an additional intramolecular cross-link.


Abstract

The conformational stability and flexibility of insulin containing a cross-link between the alpha-amino group of the A-chain to the epsilon-amino group of Lys29 of the B-chain was examined. The cross-link varied in length from 2 to 12 carbon atoms. The conformational stability was determined by guanidine hydrochloride-induced equilibrium denaturation and flexibility was assessed by H2O/D2O amide exchange. The cross-link has substantial effects on both conformational stability and flexibility which depend on its length. In general, the addition of a cross-link enhances conformational stability and decreases flexibility. The optimal length for enhanced stability and decreased flexibility was the 6-carbon link. For the 6-carbon link the Gibbs free energy of unfolding was 8.0 kcal/mol compared to 4.5 kcal/mol for insulin, and the amide exchange rate decreased by at least 3-fold. A very short cross-link (i.e. the 2-carbon link) caused conformational strain that was detectable by a lack of stabilization in the Gibbs free energy of unfolding and enhancement in the amide exchange rate compared to insulin. The effect of the cross-link length on insulin hydrodynamic properties is discussed relative to previously obtained receptor binding results. Study holds ProTherm entries: 5157 Extra Details: additive : EDTA(1 mM), conformational stability; flexibility; H2O/D2O amide exchange;,hydrodynamic properties; receptor binding

Submission Details

ID: 8t5hLSwC

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:28 p.m.

Version: 1

Publication Details
Brems DN;Brown PL;Nakagawa SH;Tager HS,J. Biol. Chem. (1991) The conformational stability and flexibility of insulin with an additional intramolecular cross-link. PMID:1988440
Additional Information

Study Summary

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 PDB Entries

Structure ID Release Date Resolution Structure Title
1BZV 1998-11-04T00:00:00+0000 0 [D-ALAB26]-DES(B27-B30)-INSULIN-B26-AMIDE A SUPERPOTENT SINGLE-REPLACEMENT INSULIN ANALOGUE, NMR, MINIMIZED AVERAGE STRUCTURE
1BZV 1998-11-04T00:00:00+0000 0 [D-ALAB26]-DES(B27-B30)-INSULIN-B26-AMIDE A SUPERPOTENT SINGLE-REPLACEMENT INSULIN ANALOGUE, NMR, MINIMIZED AVERAGE STRUCTURE
3WS3 2014-02-28T00:00:00+0000 2.33 Crystal Structure of H-2D in complex with an insulin derived peptide
6CE7 2018-02-11T00:00:00+0000 7.4 Insulin Receptor ectodomain in complex with one insulin molecule
6CE9 2018-02-11T00:00:00+0000 4.3 Insulin Receptor ectodomain in complex with two insulin molecules
6CEB 2018-02-11T00:00:00+0000 4.7 Insulin Receptor ectodomain in complex with two insulin molecules - C1 symmetry
1APH 1992-10-30T00:00:00+0000 2.0 CONFORMATIONAL CHANGES IN CUBIC INSULIN CRYSTALS IN THE PH RANGE 7-11
1APH 1992-10-30T00:00:00+0000 2.0 CONFORMATIONAL CHANGES IN CUBIC INSULIN CRYSTALS IN THE PH RANGE 7-11
1BPH 1992-10-30T00:00:00+0000 2.0 CONFORMATIONAL CHANGES IN CUBIC INSULIN CRYSTALS IN THE PH RANGE 7-11
1BPH 1992-10-30T00:00:00+0000 2.0 CONFORMATIONAL CHANGES IN CUBIC INSULIN CRYSTALS IN THE PH RANGE 7-11

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
200.0 A,B Insulin P01315 INS_PIG
91.3 B Insulin P01338 INS2_BATSP
91.7 B Insulin P68987 INS_PETMA
91.7 B Insulin P68988 INS_LAMFL
92.6 B Insulin C0HJI7 INS1_HUSDA
92.3 B Insulin C0HJI3 INS2_KATPE
92.3 B Insulin P09476 INS_ATRSP
92.3 B Insulin Q9W7R2 INS_VERMO
92.3 B Insulin P09477 INS_PLAFE
92.3 B Insulin P04667 INS_ONCKE
92.6 B Insulin P01340 INS_KATPE
92.6 B Insulin P12707 INS2_XENLA
96.2 B Insulin C0HJI8 INS2_HUSDA
96.6 B Insulin P21563 INS_RODSP
100.0 B Insulin P81423 INS_ACIGU
93.3 B Insulin P83770 INSL_VIGUN
92.9 B Insulin P12706 INS1_XENLA
96.3 B Insulin P68243 INS_CAIMO
96.3 B Insulin P68245 INS_ANSAN
96.3 B Insulin P01333 INS_ANAPL
191.60000000000002 A,B Insulin P01323 INS2_RAT
191.60000000000002 A,B Insulin P01326 INS2_MOUSE
93.1 B Insulin P67971 INS_SAISC
93.1 B Insulin P67972 INS_AOTTR
96.4 B Insulin P69048 INS_TRASC
96.4 B Insulin P69047 INS_TRADO
96.4 B Insulin P67969 INS_STRCA
96.4 B Insulin P67968 INS_MELGA
96.4 B Insulin P67970 INS_CHICK
96.6 B Insulin P51463 INS_SELRF
190.5 A,B Insulin P18109 INS_DIDVI
195.2 A,B Insulin Q62587 INS_PSAOB
200.0 A,B Insulin Q8HXV2 INS_PONPY
200.0 A,B Insulin P30410 INS_PANTR
200.0 A,B Insulin P30406 INS_MACFA
200.0 A,B Insulin P01308 INS_HUMAN
200.0 A,B Insulin Q6YK33 INS_GORGO
190.5 A,B Insulin P01316 INS_ELEMA
200.0 A,B Insulin P30407 INS_CHLAE
188.5 A,B Insulin P01324 INS_ACOCA
100.0 B Insulin F8WCM5 INSR2_HUMAN
187.2 A,B Insulin P01320 INS_CAMDR
196.7 A,B Insulin P01311 INS_RABIT
196.7 A,B Insulin Q91XI3 INS_ICTTR
191.9 A,B Insulin P01313 INS_CRILO
100.0 B Insulin P01318 INS_SHEEP
200.0 A,B Insulin P67974 INS_PHYMC
195.2 A,B Insulin P01310 INS_HORSE
100.0 B Insulin P06306 INS_FELCA
100.0 B Insulin P01319 INS_CAPHI
200.0 A,B Insulin P01321 INS_CANLF
190.5 A,B Insulin P01317 INS_BOVIN
200.0 A,B Insulin P67973 INS_BALPH
190.5 A,B Insulin P01314 INS_BALBO
90.5 A Insulin P01327 INS_CHICH
95.2 A Insulin P01322 INS1_RAT
95.2 A Insulin P01325 INS1_MOUSE