Elimination of a misfolded folding intermediate by a single point mutation.
We present an analysis of the folding behavior of the 159-residue major birch pollen allergen Bet v 1. The protein contains a water-filled channel running through it. Consequently, the protein has a hydrophobic shell, rather than a hydrophobic core. During the folding of the protein from either the urea-, pH-, or SDS-denatured state, Bet v 1 transiently populates a partially folded intermediate state. This state appears to be misfolded, since it has to unfold at least partially to fold to the native state. The misfolded intermediate is not, however, a result of the water-filled channel in Bet v 1. The intermediate completely disappears in the mutant Tyr --> Trp120, in which the channel is still present. Tyr120 appears to behave as a "negative gatekeeper" which attenuates efficient folding. The close structural homologue, the apple allergen Mal d 1, also folds without any detectable folding intermediates. However, the position of the transition state on the reaction coordinate, which is a measure of its overall compactness relative to the denatured and native states, is reduced dramatically from ca. 0.9 in Bet v 1 to around 0.5 in Mal d 1. We suggest that this large shift in the transition state structure is partly due to different local helix propensities. Given that individual mutations can have such large effects on folding, one should not a priori expect structurally homologous proteins to fold by the same mechanism.
Mogensen
Jesper E
JE
Department of Life Sciences, Aalborg University, Denmark.
Ipsen
Henrik
H
Holm
Jens
J
Otzen
Daniel E
DE
D000485:Allergens / Q000737:chemistry* / Q000235:genetics*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D052179:Antigens, Plant; D057927:Hydrophobic and Hydrophilic Interactions; D007700:Kinetics; D008956:Models, Chemical; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry; D017354:Point Mutation; D011489:Protein Denaturation; D017510:Protein Folding; D015398:Signal Transduction / Q000235:genetics; D013050:Spectrometry, Fluorescence; D040681:Structural Homology, Protein; D013816:Thermodynamics; D014364:Tryptophan / Q000235:genetics; D014443:Tyrosine / Q000235:genetics; D014508:Urea
D000485:Allergens; D052179:Antigens, Plant; C095551:MALD1 protein, Malus domestica; D010940:Plant Proteins; D014443:Tyrosine; D014364:Tryptophan; D014508:Urea