James Gumbart, Michael C. Wiener, and Emad Tajkhorshid.
Coupling of calcium and substrate binding through loop alignment in
the outer membrane transporter BtuB.
Journal of Molecular Biology, 393:1129-1142, 2009.
(PMC: 2775145)
GUMB2009
In Gram-negative bacteria, TonB-dependent outer-membrane transporters
bind large, scarce organometallic substrates with high affinity preceding
active transport. The cobalamin transporter BtuB requires the additional
binding of two Ca2+ ions before substrate binding can occur, but the
underlying molecular mechanism is unknown. Using the crystallographic
structures available for different bound states of BtuB, we have carried out
extended molecular dynamics simulations of multiple functional states of
BtuB to address the role of Ca2+ in substrate recruitment.We find that Ca2+
binding both stabilizes and repositions key extracellular loops of BtuB,
optimizing interactions with the substrate. Interestingly, replacement by
Mg2+ abolishes this effect, in accordance with experiments. Using a set of
new force-field parameters developed for cyanocobalamin, we also
simulated the substrate-bound form of BtuB, where we observed interactions
not seen in the crystal structure between the substrate and loops
previously found to be important for binding and transport. Based on our
results, we suggest that the large size of cobalamin compared to other TonB-dependent
transporter substrates explains the requirement of Ca2+ binding
for high-affinity substrate recruitment in BtuB.
