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Mutagenesis supports water mediated recognition in the trp repressor‐operator system.
Author(s) -
Joachimiak A.,
Haran T.E.,
Sigler P.B.
Publication year - 1994
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1002/j.1460-2075.1994.tb06270.x
Subject(s) - biology , repressor , operator (biology) , mutagenesis , genetics , mutation , microbiology and biotechnology , computational biology , gene , transcription factor
High resolution crystallographic analysis of the trp repressor‐operator complex indicates that the principal determinants of specificity are water mediated hydrogen bonds between the helix‐turn‐helix and the identity elements of the operator. One such hydration site involves a conserved G‐C base pair (designated G6) six nucleotides away from the dyad which, if changed symmetrically to any other pair (e.g. G6‐‐>A) reduces affinity to nonspecific levels. This same water site also contacts the conserved A5 which, if changed to G (mutation A5‐‐>G), also diminishes affinity. The stereochemistry of the water mediated hydrogen bonding system predicts that the severe deterioration of in vitro binding caused by G6‐‐>A should be reverted by a second deleterious mutation A5‐‐>G. This proved to be the case. No other second mutation at conserved operator position 5 or 7 (flanking the G6‐‐>A) reversed the effect of G6‐‐>A.