Reference : Impact of the native-state stability of human lysozyme variants on protein secretion ...
Scientific journals : Article
Life sciences : Biochemistry, biophysics & molecular biology
http://hdl.handle.net/2268/22328
Impact of the native-state stability of human lysozyme variants on protein secretion by Pichia pastoris
English
Kumita, Janet [ > > ]
Johnson, Russel [ > > ]
Alcocer, Marcos [ > > ]
Dumoulin, Mireille mailto [Université de Liège - ULg > Département des sciences de la vie > Enzymologie et repliement des protéines >]
Holmqvist, Fredrik [ > > ]
McCammon, Margaret [ > > ]
Robinson, Carol [ > > ]
Archer, David [ > > ]
Dobson, Christopher M. [ > > ]
2006
FEBS Journal
Blackwell Publishing
273
711-720
Yes (verified by ORBi)
International
1742-464X
Oxford
United Kingdom
[en] We report the secreted expression by Pichia pastoris of two human lysozyme
variants F57I and W64R, associated with systemic amyloid disease,
and describe their characterization by biophysical methods. Both variants
have a substantially decreased thermostability compared with wild-type
human lysozyme, a finding that suggests an explanation for their increased
propensity to form fibrillar aggregates and generate disease. The secreted
yields of the F57I and W64R variants from P. pastoris are 200- and 30-fold
lower, respectively, than that of wild-type human lysozyme. More comprehensive
analysis of the secretion levels of 10 lysozyme variants shows that
the low yields of these secreted proteins, under controlled conditions, can
be directly correlated with a reduction in the thermostability of their native
states. Analysis of mRNA levels in this selection of variants suggests that
the lower levels of secretion are due to post-transcriptional processes, and
that the reduction in secreted protein is a result of degradation of partially
folded or misfolded protein via the yeast quality control system. Importantly,
our results show that the human disease-associated mutations do not
have levels of expression that are out of line with destabilizing mutations
at other sites. These findings indicate that a complex interplay between
reduced native-state stability, lower secretion levels, and protein aggregation
propensity influences the types of mutation that give rise to familial
forms of amyloid disease.
http://hdl.handle.net/2268/22328

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