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Native molecular-mass estimation for PEPC was performed during FPLC on the Superdex-200 column, as described above.
Protein molecular mass was estimated as previously described.
A 500 ml column packed with Sephadex G-200 resin (pre-equilibrated with 0.1 M Tris-HCl buffer pH 8.0 containing 0.1 M NaCl) was used for the estimation of molecular mass of the affinity-purified protein.
Comparison of the elution volume of the major peak with that of protein standards of known size allows an estimation of the molecular mass of the BmrC/BmrD detergent complex of ∼310 kDa (more precisely the Stokes' radius RS is ∼6.1 nm).
Molecular mass of the protein bands was estimated using the 198 6.8 kDa molecular mass protein standards (Bio-Rad Laboratories, California, USA).
The full scan cutoff at m/z 350 was set based on the estimation that a change in the molecular mass > + 80 u was unlikely for an OTP of E1.
Furthermore, molecular weight estimation for SpGH38 by size exclusion chromatography indicated a molecular mass of 250 kDa (data not shown).
The relative molecular masses were estimated by comparisons with protein standard molecular mass markers.
M Molecular mass standards.
Kilodalton (molecular mass).
Dalton (molecular mass).
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