Philippe Rondard, Hugues Bedouelle
Mutational Scanning of a Hairpin Loop in the Tryptophan Synthase ?-Subunit Implicated in Allostery and Substrate Channeling
The tryptophan synthases from Escherichia coli and
Salmonella typhimurium are tetrameric enzymes, with
an elongated TrpA.TrpB.TrpB.TrpA structure. Structural
studies have identified residues 277–283 of TrpB
as a potentially important region for the allosteric
communication between the TrpA and TrpB subunits
and for the transport of indole between their active
sites through a hydrophobic tunnel. To explore the
functional role of this region, we analyzed the effects
of 19 single and double mutations in TrpB on the
tryptophan synthase (TSase) and serine deaminase
(SDase) activities of the TrpB2 dimer, either in the presence
or in the absence of the TrpA subunit. The mutations
of residues 273–283 could be divided into
4 classes. Mutations I278A, F280G and M282A decreased
the SDase and TSase activities of TrpB2 to
similar extents. F280A decreased the SDase activity
of TrpB2 more than its TSase activity, whereas the reverse
was true for Y279L. F280A decreased the activation
factor of TrpB2 by TrpA, whereas F280G increased
it. The reaction steps and intramolecular contacts that
could be affected by the mutations are described. The
sequence 278-IYFGM-282, which is present in E. coli
and S. typhimurium, is only found in 5 out of 42 organisms,
whereas the sequence VLHGX is found in 21 organisms.
Our results identified several mutations that
could be used as structural probes to analyze precisely
the roles of residues 278–282 and their evolution.
Biological Chemistry, Walter de Gruyter
Print ISSN: 1431-6730
Volume: 381, 12/2000
Pages: 1185 - 1193
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