Quantifying In Situ Structural Stabilities of Human Blood Plasma Proteins Using a Novel Iodination Protein Stability Assay

Many of the diseases
that plague society today are driven by a
loss of protein quality. One method to quantify protein quality is
to measure the protein folding stability (PFS). Here, we present a
novel mass spectrometry (MS)-based approach for PFS measurement, iodination
protein stability assay (IPSA). IPSA quantifies the PFS by tracking
the surface-accessibility differences of tyrosine, histidine, methionine,
and cysteine under denaturing conditions. Relative to current methods,
IPSA increases protein coverage and granularity to track the PFS changes
of a protein along its sequence. To our knowledge, this study is the
first time the PFS of human serum proteins has been measured in the
context of the blood serum (in situ). We show that IPSA can quantify
the PFS differences between different transferrin iron-binding states
in near in vivo conditions. We also show that the
direction of the denaturation curve reflects the in vivo surface accessibility of the amino acid residue and reproducibly
reports a residue-specific PFS. Along with IPSA, we introduce an analysis
tool Chalf that provides a simple workflow to calculate
the residue-specific PFS. The introduction of IPSA increases the potential
to use protein structural stability as a structural quality metric
in understanding the etiology and progression of human disease. Data
is openly available at Chorusproject.org (project ID 1771).