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six peaks, the change in contour length between consecutive peaks was 8.4
nm, which corresponds to the lengthening of the 36 amino acids of a single
repeat. Urea strongly altered the shape of the unfolding peaks, conirming
that disruption of the protein hydrogen bonds leads to a loss of mechanical
stability. This observation correlates with the cellular behaviour since Als5p-
mediated adhesion has been shown to be reversibly inhibited by urea and
formamide.
(c)
(a)
(b)
Figure 15.6.
Unfolding single cell adhesion proteins. (a) Als5p contains a tandem
repeat (TR) region comprising multiple glycosylated 36-amino acid repeats that
are arranged in anti-parallel
B
-sheets. (b) Ig-T-TR
6
fragments were attached on a
gold surface and stretched via their Ig domains using an Ig-T tip. (c) Force-extension
curves obtained by stretching single Ig-T-TR
6
showed periodic features relecting
the sequential unfolding of the TR domains (upper traces). Force peaks were well
described by the WLC model (inset; red line). Addition of urea dramatically altered
the unfolding peaks (lower traces). Reprinted with permission from Alsteens
et al.
40
Remarkably, single Als proteins could also be unfolded on live cells. Force
curves obtained on yeast cells expressing six repeats displayed sawtooth
patterns similar to those found on isolated proteins, while cells expressing no
repeat were unable to bind the AFM tip. The unfolding probability increased
with the number of repeats and was correlated with the level of cell-cell
adhesion, indicating these modular domains may play a role in fungal
adhesion. The modular and lexible nature of Als conveys both strength and
toughness to the protein, making it ideally suited for cell adhesion. These
single-molecule measurements provide novel insights into the mechanical
properties of adhesion molecules and may help us to elucidate their potential
implication in diseases.
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