Localized nature of the transition-state structure in goat alpha-lactalbumin folding.


To investigate whether the structure partially formed in the molten globule folding intermediate of goat alpha-lactalbumin is further organized in the transition state of folding, we constructed a number of mutant proteins and performed Phi-value analysis on them. For this purpose, we measured the equilibrium unfolding transitions and kinetic refolding and unfolding reactions of the mutants using equilibrium and stopped-flow kinetic circular dichroism techniques. The results show that the mutants with mutations located in the A-helix (V8A, L12A), the B-helix (V27A), the beta-domain (L52A, W60A), the C-helix (K93A, L96A), the C-D loop (Y103F), the D-helix (L105A, L110A), and the C-terminal 3(10)-helix (W118F), have low Phi-values, less than 0.2. On the other hand, D87N, which is located on the Ca(2+)-binding site, has a high Phi-value, 0.91, indicating that tight packing of the side-chain around Asp87 occurs in the transition state. One beta-domain mutant (I55V) and three C-helix mutants (I89V, V90A, and I95V) demonstrated intermediate Phi-values, between 0.4 and 0.7. These results indicate that the folding nucleus in the transition state of goat alpha-LA is not extensively distributed over the alpha-domain of the protein, but very localized in a region that contains the Ca(2+)-binding site and the interface between the C-helix and the beta-domain. This is apparently in contrast with the fact that the molten globule state of alpha-lactalbumin has a partially formed structure inside the alpha-domain. It is concluded that the specific docking of the alpha and beta-domains at a domain interface is necessary for this protein to organize its native structure from the molten globule intermediate. Study holds ProTherm entries: 17261, 17262, 17263, 17264, 17265, 17266, 17267, 17268, 17269, 17270, 17271, 17272, 17273, 17274, 17275, 17276, 17277, 17278, 17279, 17280, 17281, 17282, 17283, 17284, 17285, 17286, 17287, 17288, 17289, 17290, 17291, 17292, 17293, 17294, 17295, 17296, 17297, 17298, 17299, 17300, 17301, 17302, 17303, 17304, 17305, 17306, 17307, 17308 Extra Details: protein folding; alpha-lactalbumin; the transition state; the molten globule state; phi-value analysis

Submission Details

ID: myRPya2C

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:49 p.m.

Version: 1

Publication Details
Saeki K;Arai M;Yoda T;Nakao M;Kuwajima K,J. Mol. Biol. (2004) Localized nature of the transition-state structure in goat alpha-lactalbumin folding. PMID:15276846
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 Alpha-lactalbumin P00712 LALBA_CAPHI
97.9 Alpha-lactalbumin P09462 LALBA_SHEEP
96.5 Alpha-lactalbumin Q9TSN6 LALBA_BUBBU
95.1 Alpha-lactalbumin P00711 LALBA_BOVIN
94.4 Alpha-lactalbumin Q9TSR4 LALBA_BOSMU