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Altering ionic
strength and pH changes the folding pathway of lysozyme
Sunita Kulkarni, Department of
Biochemistry and Molecular Biology,
University of Leeds Woodhouse Lane, Leeds West Yorkshire, LS2.
9JT.
UK
Hen egg white lysozyme has been widely used as a model system for
folding studies as it is small, has a well characterised
structure,
and exhibits reversible folding over a measurable time scale.
A similar multipronged biophysical approach as that used for
elucidating the protein folding pathway of lysozyme at pH 5.2 and
20'C (Radford and Dobson, 1995) has been employed to analyse the
pathway under physiological conditions. In summary, at pH 5.2,
hydrophobic collapse occurs within 10ms, followed by the
formation
of stable a-helical structure after a further 50ms. Finally on a
300ms time scale, the native state of the enzyme forms with
correct
a and b domains. 75% of lysozyme molecules are thought to follow
this pathway, but the remaining 25% may undergo a faster folding
mechanism with a rate on a 100ms time scale to get to the native
state. We are dissecting the kinetic and thermodynamic barriers
thought to be responsible for the generation of the parallel
pathways
described above. We show here, that pH and ionic strength are
important factors in the alteration of the folding mechanism from
that observed at pH 5.2, leading to changes in the rate limiting
step of folding, and to a sequential single pathway.The ways in
which
electrostatics, ionisation and hydrophobic effects may influence
the
early stages of protein folding, leading to routing along
'folding
funnels' to the native state are considered.
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