Abstract:
:To study fibronectin (FN) conformation and assembly, we generated several deletion mutants: FNΔI1-5, FNΔIII1-3, FNΔIII4-8, and FNΔIII11-14. A monomeric form, FNmono, which lacked the C-terminal dimerization region, was also created. FNtnA-D was generated by swapping FNIII domains 1-8 in FNΔIII11-14 with seven FNIII domains from tenascin-C. The conformations of these mutants were analyzed by glycerol gradient sedimentation under low-salt (20 mM NaCl) and high-salt (200 mM NaCl) conditions. Surprisingly, most of the mutants showed a compact conformation under low-salt conditions, except for FNtnA-D. When we tested these mutants in cell culture, FNΔI1-5, FNΔIII1-3, and FNtnA-D were unable to form a matrix. Interestingly, FNΔIII1-3 and FNtnA-D were capable of co-assembly with full-length FN, while FNΔI1-5 was not. This indicates that the segment I1-5 is crucial for matrix assembly and segment III1-3 is also important. Mutations in FN are associated with glomerulopathy, but when we studied mutant proteins, the single-nucleotide mutations had only minor effects on conformation and matrix assembly. The mutations may destabilize their FNIII domains or generate dimers of dimers by disulfide cross-linking.
journal_name
Biochemistryjournal_title
Biochemistryauthors
Ohashi T,Lemmon CA,Erickson HPdoi
10.1021/acs.biochem.7b00589subject
Has Abstractpub_date
2017-08-29 00:00:00pages
4584-4591issue
34eissn
0006-2960issn
1520-4995journal_volume
56pub_type
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