Two-step folding of recombinant mitochondrial porin in detergent.

Abstract:

:Precise information regarding the transmembrane topology of mitochondrial porin is essential for understanding the mechanisms by which this protein functions. Porin acts as a channel in the outer membrane and interacts with small solutes and proteins to regulate mitochondrial function. The acquisition of high-resolution structural data requires a method of maintaining high concentrations of unaggregated, properly folded porin. In the current studies, several mixed detergent systems were analyzed for their ability to fold Neurospora mitochondrial porin expressed in and isolated from Escherichia coli. A mixture of sodium dodecyl sulfate and dodecyl-beta-D-maltopyranoside in a 1:6 molar ratio supports a beta-strand-rich conformation. In this state, the two tryptophan residues in the protein reside in hydrophobic environments, and about half of the nine tyrosines are solvent exposed. Most importantly, heat-labile tertiary contacts, as detected by near-UV circular dichroism spectropolarimetry, in the sodium dodecyl sulfate/dodecyl-beta-D-maltopyranoside-solubilized porin are very similar to those of the protein following functional reconstitution into liposomes. Similarly, both forms are protease resistant. Thus, a method has been identified with the potential to solubilize high concentrations of mitochondrial porin in a state virtually indistinguishable from the membrane-embedded form.

journal_name

Biophys J

journal_title

Biophysical journal

authors

Bay DC,O'Neil JD,Court DA

doi

10.1529/biophysj.107.115196

subject

Has Abstract

pub_date

2008-01-15 00:00:00

pages

457-68

issue

2

eissn

0006-3495

issn

1542-0086

pii

S0006-3495(08)70723-9

journal_volume

94

pub_type

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