Pathway of oxidative folding of a 3-disulfide alpha-lactalbumin may resemble either BPTI model or hirudin model.
Pathways of oxidative folding of disulfide proteins display a high degree of diversity and vary among two extreme models. The BPTI model is defined by limited species of folding intermediates adopting mainly native disulfide bonds. The hirudin model is characterized by highly heterogeneous folding intermediates containing mostly non-native disulfide bonds. alphaLA-IIIA is a 3-disulfide variant of alpha-lactalbumin (alphaLA) with a 3-D conformation essentially identical to that of intact alphaLA. alphaLA-IIIA contains 3 native disulfide bonds of alphaLA, two of them are located at the calcium binding beta-subdomain (Cys61-Cys77 and Cys73-Cys91) and the third bridge is located within the alpha-helical domain of the molecule (Cys28-Cys111). We investigate here the pathway of oxidative folding of fully reduced alphaLA-IIIA with and without stabilization of its beta-subdomain by calcium binding. In the absence of calcium, the folding pathway of alphaLA-IIIA was shown to resemble that of hirudin model. Upon stabilization of beta-sheet domain by calcium binding, the folding pathway of alphaLA-IIIA exhibits a striking similarity to that of BPTI model. Three predominant folding intermediates of alphaLA-IIIA containing exclusively native disulfide bonds were isolated and structurally characterized. Our results further demonstrate that stabilization of subdomains in a protein may dictate its folding pathway and represent a major cause for the existing diversity in the folding pathways of the disulfide-containing proteins.
Salamanca
Silvia
S
Research Center for Protein Chemistry, Institute of Molecular Medicine, 2121 W. Holcombe Blvd., Houston, TX 77030, USA.
Chang
Jui-Yoa
JY
D000595:Amino Acid Sequence; D000818:Animals; D007611:Aprotinin / Q000737:chemistry* / Q000378:metabolism; D002118:Calcium / Q000494:pharmacology; D002417:Cattle; D004220:Disulfides / Q000737:chemistry*; D006629:Hirudins / Q000737:chemistry* / Q000378:metabolism; D007768:Lactalbumin / Q000737:chemistry* / Q000378:metabolism*; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction / Q000187:drug effects; D017510:Protein Folding
D004220:Disulfides; D006629:Hirudins; D007768:Lactalbumin; D007611:Aprotinin; D002118:Calcium