Ms. Kindy et al., ADENOVIRAL EXPRESSION OF MURINE SERUM AMYLOID-A PROTEINS TO STUDY AMYLOID FIBRILLOGENESIS, Biochemical journal, 332, 1998, pp. 721-728
Serum amyloid A (SAA) proteins are one of the most inducible acute-pha
se reactants and are precursors of secondary amyloidosis. In the mouse
, SAA(1) and SAA(2) are induced in approximately equal quantities in r
esponse to amyloid induction models. These two isotypes differ in only
9 of 103 amino acid residues; however, only SAA(2) is selectively dep
osited into amyloid fibrils. SAA expression in the CE/J mouse species
is an exception in that gene duplication did not occur and the CE/J va
riant is a hybrid molecule sharing features of SAA(1) and SAA(2). Howe
ver, even though it is more closely related to SAA(2) it is not deposi
ted as amyloid fibrils, We have developed an adenoviral vector system
to overexpress SAA proteins in cell culture to determine the ability o
f these proteins to form amyloid fibrils, and to study the structural
features in relation to amyloid formation. Both the SAA(2) and CE/J SA
A proteins were synthesized in large quantities and purified to homoge
neity. Electron microscopic analysis of the SAA proteins revealed that
the SAA(2) protein was capable of forming amyloid fibrils, whereas th
e CE/J SAA was incapable. Radiolabelled SAAs were associated with norm
al or acute-phase high-density lipoproteins (HDLs); we examined them f
or their clearance from the circulation. In normal mice, SAA(2) had a
half-life of 70 min and CE/J SAA had a half-life of 120 min; however,
in amyloid mice 50 % of the SAA(2) cleared in 55 min, compared with 13
5 min for the CE/J protein. When the SAA proteins were associated with
acute-phase HDLs, SAA(2) clearance was decreased to 60 min in normal
mice compared with 30 min in amyloidogenic mice. Both normal and acute
-phase HDLs were capable of depositing SAA(2) into preformed amyloid f
ibrils, whereas the CE/J protein did not become associated with amyloi
d fibrils. This established approach opens the doors for large-scale S
AA production and for the examination of specific amino acids involved
in the fibrillogenic capability of the SAA(2) molecule in vitro and i
n vivo.