TY - JOUR
T1 - Interdomain Communication between the Thiolation and Thioesterase Domains of EntF Explored by Combinatorial Mutagenesis and Selection
AU - Zhou, Zhe
AU - Lai, Jonathan R.
AU - Walsh, Christopher T T.
N1 - Funding Information:
This research was supported by the National Institutes of Health (AI042738, GM020011, and GM047467). J.R.L. is supported by a post-doctoral fellowship from the Helen Hay Whitney Foundation. We thank Michael Fischbach, Ellen Yeh, and Hening Lin for helpful discussion and critical reading of this manuscript.
PY - 2006/8
Y1 - 2006/8
N2 - Thiolation (T) domains are protein way stations in natural product assembly lines. In the enterobactin synthetase, the T domain on EntF is recognized in cis by its catalytic partners: the EntF condensation (C), adenylation (A), and thioesterase (TE) domains. To assess surface features of the EntF T domain recognized by C, A, and TE, regions of the EntF T domain were submitted to shotgun alanine scanning and Ent production selection, which revealed residues that could not be substituted by Ala. EntF mutants bearing Ala in such positions were assayed in vitro for Ent production with EntEB, and for A-T, C-T, and T-TE communications. We concluded that G1027A and M1030A are specifically defective in acyl transfer from T to TE. These residues define an interaction surface between these two in cis domains in an NRPS module.
AB - Thiolation (T) domains are protein way stations in natural product assembly lines. In the enterobactin synthetase, the T domain on EntF is recognized in cis by its catalytic partners: the EntF condensation (C), adenylation (A), and thioesterase (TE) domains. To assess surface features of the EntF T domain recognized by C, A, and TE, regions of the EntF T domain were submitted to shotgun alanine scanning and Ent production selection, which revealed residues that could not be substituted by Ala. EntF mutants bearing Ala in such positions were assayed in vitro for Ent production with EntEB, and for A-T, C-T, and T-TE communications. We concluded that G1027A and M1030A are specifically defective in acyl transfer from T to TE. These residues define an interaction surface between these two in cis domains in an NRPS module.
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U2 - 10.1016/j.chembiol.2006.06.011
DO - 10.1016/j.chembiol.2006.06.011
M3 - Article
C2 - 16931336
AN - SCOPUS:33747374631
SN - 1074-5521
VL - 13
SP - 869
EP - 879
JO - Chemistry and Biology
JF - Chemistry and Biology
IS - 8
ER -