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Papers of the Month - September 1997
ANEURYSM PAPERS OF THE MONTH - SEPTEMBER 97
Two papers have recently come out from our laboratory, describing
three more possible members of the family of aortic aneurysm
antigenic proteins (AAAP's).
1. Hirose H, Ozsvath KJ, Xia S, Tilson MD. Molecular cloning of
the cDNA for an additional member of the family of aorftic aneurysm
antigenic proteins. J Vasc Surg 1997; 26: 313-8.
Purpose: We have purified and partially sequenced a protein from
the adventitia of the human aorta (AAAP-40) that has homologies to
bovine aorta-specific MAGP-36. AAAP-40 and MAGP-36 have fibrinogen
(FBN)-like and vitronectin (VN)-like motifs. Screening an aortic
adventitia expression library with antibodies against FBN and VN
has resulted in the cloning of three cDNA's, the recombinant
proteins of which are immunoreactive with IgG from patients with
AAA. Two strongly resemble each other and are described in the
next abstract. The present abstract reports the third, here called
clone 4.
Methods: mRNA from a specimen of human AAA adventitia was reverse-
transcribed for insertion into the phagemid UniZap MFR'
(Stratagene). The library was transfected to E Coli and screened
with rabbit antihuman VN antibody.
Results: The hypothetical protein encoded by clone 4 shares
sequence motifs with known microfibril-associated glygoproteins
(MAGP's). There were also similarities to members of the
immunoglobulin kappa family, to fibrinogen-beta, and to
cytomegalovirus (which is a potential molecular mimic).
Recombinant clone 4 was immunoreactive with serum from three of
four patients with AAA and has a MW of approximately 28 kDa.
Conclusion: There may be several members of the AAAP family.
2. Ozsvath KJ, Hirose H, Xia S, Chew D, Knoetgen J, Tilson MD.
Expression of two novel recombinant proteins from aortic adventitia
(kappafibs) sharing amino acid sequences with cytomegalovirus. J
Surgical Research (May 1997), volume 69.
Abstract: AAAP-40, in addition to the features described in the
above abstract, is immunoreactive with IgG's purified from the
surgical specimens of patients undergoing AAA repair, and also from
their serum. In addition to similarities to VN and the FBN's, it
shares motifs with IgKappa sequences (which may be binding sites
for integrins) and cytomegalovirus (which may be a molecular mimic
in the pathogenesis of AAA). This communication describes the
expression of two additional clones, with lengthy IgKappa-like
sequences (~100 residues), in a strain of E Coli. The expressed
proteins were separated by SDS/PAGE, and the immunoblots were
probed with AAA IgG. Experimental cell lines, transfected with the
clones (clones 1 and 5), synthesized recombinant proteins,
detectable in Western immunoblots with AAA IgG at a MW of
approximately 28 kDa. A prediction of the tertiary structure of
the protein encoded by clone 1 resembles the well-characterized
immunoglobulin fold of the cell adhesion molecules. Pairwise
sequence analysis also revealed significant similarities to a
protein of cytomegalovirus. A ClustalW multiple sequence alignment
is presented, which illustrates the similarities of eight proteins:
three aneurysm antigenic proteins, two Ig kappas, MAGP-36, MFAP-4,
and a fibrinogen-like peptide of the invertebrate echinoderm.
To examine a novel hypothesis that matrix cell adhesion molecules
carry out a more primitive function than soluble immunity (which
did not appear until the cartilagenous sharks), we used AllAll to
calculate the point-accepted mutation rates (PAM units) for several
of the above-mentioned eight proteins, plus VCAM-1 (human) and
fibrinogen-beta. For fibrinopeptide-related proteins, a PAM unit
has been estimated to be about 1 million years. The resulting
phylogenetic tree indicates that clone 1 is significantly closer to
an Ig kappa than any other protein under consideration, including
VCAM-1. There also appear to have been two clusters of
evolutionary activity. The first was around the time of the
Cambrian radiation (producing fibrinogen and the MAGP's; the second
was much more recent, spawning the CAMs first, and then the soluble
immunoglobulins.
The findings suggest that there may be a novel family of matrix
proteins that use immunoglobulin motifs, possibly as binding sites
for integrins; and that there are matrix proteins in addition to
AAAP-40 that may serve as autoantigens in the pathogenesis of AAA
disease.