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dc.contributor.authorBarzak, Fareeda Maged
dc.date.accessioned2015-03-05T20:30:58Z
dc.date.available2015-03-05T20:30:58Z
dc.date.issued2014
dc.identifier.urihttp://hdl.handle.net/10179/6328
dc.description.abstractThe cytoskeleton network allows cells to differentiate, divide, and move in response to the external environment creating a mechanoprotection system against cell stress. The actin cytoskeleton is stabilised and tightly regulated by various actin-binding proteins, one of which are the family of Filamin (FLN) proteins that crosslink F-actin into three-dimensional networks. Filamins also link the actin cytoskeleton to the cellular membrane through interactions with transmembrane proteins and function as a molecular scaffold for signalling molecules. In addition to an actin binding domain, each monomer contains a rod region of 24 immunoglobulin-like repeat domains with dimerisation of the monomers occurring at repeat 24. The human filamin family contains three FLN isoforms; FLNA, FLNB, and FLNC which are differentially expressed where FLNA is identified as the dominant isoform located on the X-chromosome essential for mammalian development. Mutations in Filamin A (FLNA) have been identified to cause distinctly different human diseases affecting the central nervous system, vascular system, or skeletal muscles; however, the molecular mechanisms of FLNA leading to these diseases remain unclear. Mutations cluster in distinct FLNA domains, suggesting their functional importance for mediating correct functions. Mutations in the FLNA repeat 10 domain are correlated with severe forms of the skeletal disorders Otopalatodigital syndrome spectrum disorders (OPD) thought to be due to an altered or gain-of-function phenotype. The aim of this study was to provide an insight into the biochemical properties of FLNA repeat 10 domain by better understanding how mutations in this domain lead to OPD. Initially, recombinant wildtype (Wt) and mutant (V1249A and A1188T) FLNA repeat 10 domain proteins (FLNAR10) were purified then compared by in vitro biochemical studies to investigate secondary structure, stability, and affinity towards F-actin. The FLNAR10 protein was revealed to have relatively weak binding affinity towards F-actin, consistent with being an additional contributor in the filamin protein to bind F-actin. Mutations in the FLNAR10 protein exhibited a slight increase in affinity towards F-actin, accompanied by a slight reduction of thermostability in comparison to the Wt protein, but no significant changes in the secondary structure were observed. This slight increase in the affinity of the mutant FLNA repeat 10 proteins towards F-actin is consistent with a gain-of function mechanism for the disease phenotype. Overall, these results contribute towards a better understanding of the FLNA function, providing further evidence towards a gain-of function mechanism for OPD.en_US
dc.language.isoenen_US
dc.publisherMassey Universityen_US
dc.rightsThe Authoren_US
dc.subjectMicrofilament proteinsen_US
dc.subjectSkeletal disordersen_US
dc.subjectFilamin proteinsen_US
dc.subjectActin-cytoskeletonen_US
dc.subjectOtopalatodigital syndromeen_US
dc.titleHow does the interaction between the Filamin A repeat 10 domain and F-actin lead to severe OPD skeletal disorders? : a thesis presented in partial fulfilment of the requirements for the degree of Master of Science in Biochemistry at Massey University, Palmerston North, New Zealanden_US
dc.typeThesisen_US
thesis.degree.disciplineBiochemistryen_US
thesis.degree.grantorMassey Universityen_US
thesis.degree.levelMastersen_US
thesis.degree.nameMaster of Science (M.Sc.)en_US


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