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Folding of T1 subdomains of nascent Kv1.3
Author(s) -
Kosolapov Andrey,
Deutsch Carol
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.5.a965-c
Subject(s) - folding (dsp implementation) , linker , chemistry , c terminus , biogenesis , n terminus , biophysics , crystallography , protein folding , amino acid , stereochemistry , peptide sequence , biochemistry , biology , computer science , electrical engineering , engineering , gene , operating system
Kv channels contain a recognition domain, T1, in the amino terminus that facilitates Kv tetramerization. We have previously determined when and where global folding of this domain occurs ( Kosolapov et al., Neuron 44 : 296 – 307 , 2004 ; Kosolapov and Deutsch , J. Biol. Chem. 278 : 4305 – 4313 , 2003 ). We now ask two questions: what is the elemental unit of folding inside the ribosomal tunnel? Do T1 subdomains fold in a sequential and independent fashion or in a concerted manner? Two subdomains were investigated: a helical hairpin in the C‐terminus of T1 and an anti‐parallel β‐strand in the N‐terminus. Using Kv1.3 biogenic intermediates of different lengths that contain crosslinkable cysteine pairs engineered at the folded intrasubunit interface of each subdomain, we have determined the length of peptide linker at the C‐terminal end of each subdomain that permits tertiary structure formation. Our methods include intramolecular crosslinking and pegylation techniques. Neither subdomain folds efficiently until it emerges from the ribosomal exit tunnel, suggesting that the tunnel cannot accommodate these minimalist tertiary structures. Moreover, tertiary folding of subdomains appears to be independent and sequential. A plot of the probability of folding as a function of fractional accessibility permits us to calculate the relative free energies of folding for each subdomain. These studies provide insight into folding events during early Kv channel biogenesis. ([Supported by NIH grant GM 52302].