Date of Award

2016

Degree Type

Thesis

Degree Name

M.Sc.

Department

Chemistry and Biochemistry

First Advisor

Koschinsky, Marlys

Keywords

Apolipoprotein(a); Atherosclerosis; I4399M; Lipoprotein(a)

Rights

CC BY-NC-ND 4.0

Abstract

Elevated plasma levels of lipoprotein(a) (Lp(a)) are a causal risk factor for CHD. Lp(a) closely resembles LDL, but contains an additional glycoprotein apolipoprotein(a) (apo(a)) that is structurally homologous to the fibrinolytic proenzyme plasminogen. This has led to speculation that Lp(a) can oppose the fibrinolytic functions of plasminogen. A single nucleotide polymorphism (SNP) in the LPA gene encoding apo(a) results in an Ile to Met substitution at position 4399 in the protease-like domain. In population studies, this variant has been correlated with elevated plasma Lp(a) levels and with higher CHD risk. We undertook a functional characterization of the effect of the I4399M substitution in apo(a). Molecular dynamics simulations of wild-type (wt) apo(a) and the Met variant revealed a shift from a buried (Ile) to slightly exposed (Met) environment. Subsequent MALDI-TOF mass spectrometry analysis demonstrated the presence of a methionine sulfoxide moiety at this position in the Met variant. When 17-kringle recombinant forms of apo(a) were included in a plasma clot lysis assay, both the wt apo(a) and Met variant inhibited lysis, but the Met variant had a 50% greater effect. However, the Met variant was equally as efficient as wt apo(a) in inhibiting plasminogen activation on a fibrin surface. The Met variant was also able to significantly shorten coagulation time and result in greater turbidity for clots made from either purified fibrin or lipoprotein-deficient plasma compared to wild-type. In agreement with these findings, SEM and confocal microscopy of clots made from fibrin and citrated plasma showed that compared to wt apo(a), the Met variant resulted in significant alteration of the fibrin network. Together, our data suggest that the Met4399 variant differs structurally from wt apo(a), which may underlie key differences related to its effects on fibrin clot architecture and fibrinolysis.

Available for download on Tuesday, February 27, 2018

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