TrpB2 Enzymes are O-Phospho-l-serine Dependent Tryptophan Synthases

Abstract

ABSTRACT:The rapid increase of the number of sequenced genomes asks for the functional annotation of the encoded enzymes. We used a combined computational−structural approach to determine the function of the TrpB2 subgroup of the tryptophan synthase β chain/β chain-like TrpB1−TrpB2 family (IPR023026). The results showed that TrpB2 enzymes are O-phospho-L-serine dependent tryptophan synthases, whereas TrpB1 enzymes catalyze the L-serine dependent synthesis of tryptophan. We found a single residue being responsible for the different substrate specificities of TrpB1 and TrpB2 and confirmed this finding by mutagenesis studies and crystallographic analysis of a TrpB2 enzyme with bound O-phospho-Lserine.

Transposon mutagenesis in Pseudomonas fluorescens reveals genes involved in blue pigment production and antioxidant protection.

“…This result confirms previous hypothesis that indicated the involvement of the second copies of trp genes in the blue pigment biosynthesis through an indole/indigo biosynthesis pathway (Andreani et al, 2015). Carriage of multiple trpB gene copies has been reported in several bacterial and archaeal species (Xie et al, 2001;Merkl, 2007;Busch et al, 2014) and are frequently involved in biosynthesis of molecules with different functions, despite the high-energy cost required for this pathway. It has been demonstrated that usually the second copy possesses substrate specificity, not assembled with TrpA and seems to have other functions, such as indole salvage (Busch et al, 2014;Hiyama et al, 2014).…”

“…Carriage of multiple trpB gene copies has been reported in several bacterial and archaeal species (Xie et al, 2001;Merkl, 2007;Busch et al, 2014) and are frequently involved in biosynthesis of molecules with different functions, despite the high-energy cost required for this pathway. It has been demonstrated that usually the second copy possesses substrate specificity, not assembled with TrpA and seems to have other functions, such as indole salvage (Busch et al, 2014;Hiyama et al, 2014). In Pseudomonas aeruginosa the first enzyme of the tryptophan biosynthesis pathway (Antranilate synthase coded by trpE and trpG genes) is duplicated and the second copy is involved in the quorum sensing signalling but not in tryptophan biosynthesis (Palmer et al, 2013).…”

Ancestral Tryptophan Synthase Reveals Functional Sophistication of Primordial Enzyme Complexes

“…Other archaeal phyla were not included as they contain a different type of TS b subunit (Busch et al, 2014;Merkl, 2007). A maximum-likelihood tree was created based on the most probable substitution model (Figure S1).…”

Light-Regulation of Tryptophan Synthase by Combining Protein Design and Enzymology

“…Expression strains used for production of TrpA, TrpB, ec TrpA, and ec TrpB, as well as of auxiliary enzymes VioA and GAP dehydrogenase, were purchased from Agilent Technologies [ E. coli BL21 Gold (DE3), BL21-CodonPlus (DE3)-RIPL] (Santa Clara, CA, USA) and Novagen [ E. coli BL21 (DE3) Rosetta] (Merck, Darmstadt, Germany). The following expression vectors ware taken from previously published work: pET24a_TrpB for TrpB expression [67], and pET28a_tmGAPDH for GAP dehydrogenase expression [68]. The plasmid pEVOL_ONBY for incorporation of ONBY into proteins was provided by Prof. Peter Schultz (Scripps Research Institute, La Jolla, CA, USA) [14,69].…”