Crystal Structures of a New Class of Allosteric Effectors Complexed to Tryptophan Synthase

Weyand, Michael; Schlichting, Ilme; Mozzarelli, Andrea

doi:10.1074/jbc.m111285200

Cited by 44 publications

(57 citation statements)

References 43 publications

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“…The second catalytically important amino acid ␣Glu 49 is modeled in two conformations and forms a hydrogen bond with the IAG acetyl oxygen atom. The implications of this interaction for the ␣-reaction are discussed in the accompanying paper (24). The hydrogen bonding patterns and distances are shown in Fig.…”

Section: Iag Structurementioning

confidence: 95%

“…IAG binds to the mutant in the same manner as to the wild-type (24), but loop ␣L6 is not closed. The indole nitrogen forms a hydrogen bond with aspartate ␣Asp 60 and the acetyl carboxylate group mimics the IGP/IPP/GP phosphate group.…”

Section: Iag Structurementioning

confidence: 99%

“…Therefore, the steady kinetics of the forward and reverse ␣-reaction were analyzed in detail. The crystal structures of wild-type tryptophan synthase complexed with IAG and other novel ␣-subunit ligands are reported in the accompanying paper (24 …”

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confidence: 99%

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Crystal Structure of the βSer178 → Pro Mutant of Tryptophan Synthase

Weyand¹,

Schlichting²,

Herde³

et al. 2002

Journal of Biological Chemistry

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The catalytic activity of the pyridoxal 5-phosphatedependent tryptophan synthase ␣ 2 ␤ 2 complex is allosterically regulated. The hydrogen bond between the helix ␤H6 residue ␤Ser 178 and the loop ␣L6 residue Gly 181 was shown to be critical in ligand-induced intersubunit signaling, with the ␣-␤ communication being completely lost in the mutant ␤Ser 178 3 Pro (Marabotti, A., De Biase, D., Tramonti, A., Bettati, S., and Mozzarelli, A. (2001) J. Biol. Chem. 276, 17747-17753). The structural basis of the impaired allosteric regulation was investigated by determining the crystal structures of the mutant ␤Ser 178 3 Pro in the absence and presence of the ␣-subunit ligands indole-3-acetylglycine and glycerol 3-phosphate. The mutation causes local and distant conformational changes especially in the ␤-subunit. The ligand-free structure exhibits larger differences at the N-terminal part of helix ␤H6, whereas the enzyme ligand complexes show differences at the C-terminal side. In contrast to the wild-type enzyme loop ␣L6 remains in an open conformation even in the presence of ␣-ligands. This effects the equilibrium between active and inactive conformations of the ␣-active site, altering k cat and K m , and forms the structural basis for the missing allosteric communication between the ␣-and ␤-subunits.The ␣ 2 ␤ 2 complex of tryptophan synthase (TRPS) 1 (EC 4.2.1.20) is a bifunctional enzyme that is considered a paradigm for the analysis of intersubunit regulatory signals. The enzyme is formed by two ␣-and ␤-subunits, arranged in a linear ␣␤␤␣ geometry (1), and catalyzes the last two steps in the biosynthesis of L-tryptophan in a highly concerted mode. The isolated ␣-and ␤-subunits exhibit an activity about 2 orders of magnitude lower that in the tetramer, indicating that the formation of the complex is accompanied by subunit conformational changes (2, 3). Further levels of regulation are achieved by ligand-induced intersubunit signals that are pH-, temperature-, and cation-dependent and keep the catalytic activities of ␣ and ␤-active sites in phase (4 -11). The mechanism of the ␣-␤ activation and allosteric regulation is based on an open-close transition of both subunits (7, 12, 13) involving the movement of several parts of the ␣-and ␤-subunits. In particular, signal transduction from the ␣-to the ␤-active site involves residues belonging to loop ␣L2, loop ␣L6, and the helix ␤H6 (14, 15), the latter structural element being part of the ␤Ϫsubunit COMM domain, as defined by Schneider et al. (15). The question whether there is a unique or multiple pathways of communication and whether these pathways are specialized in controlling defined functional properties of the enzyme was addressed by investigating the activity and regulatory properties of several mutants of the ␣-and ␤-subunits. It was found that mutants in loop ␣L2 and mutants in helix ␤H6 that interact with loop ␣L2 exhibit altered ␣ and ␤ activities (16 -18), whereas mutants in loop ␣L6 and mutants in helix ␤H6 interacting with loop ␣L6 also exhibit altered alloster...

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Section: Iag Structurementioning

confidence: 95%

Section: Iag Structurementioning

confidence: 99%

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Crystal Structure of the βSer178 → Pro Mutant of Tryptophan Synthase

Weyand¹,

Schlichting²,

Herde³

et al. 2002

Journal of Biological Chemistry

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“…Among the 21 protein family pairs, only two are known to interact based on experimentally resolved cocrystal structures: TrpA−TrpB [PDB 1k7f (34) (Fig. 4).…”

Section: Simultaneous Identification Of Interacting Families and Specmentioning

confidence: 99%

Simultaneous identification of specifically interacting paralogs and interprotein contacts by direct coupling analysis

Gueudré

Baldassi

Zamparo

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

125

173

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Understanding protein−protein interactions is central to our understanding of almost all complex biological processes. Computational tools exploiting rapidly growing genomic databases to characterize protein−protein interactions are urgently needed. Such methods should connect multiple scales from evolutionary conserved interactions between families of homologous proteins, over the identification of specifically interacting proteins in the case of multiple paralogs inside a species, down to the prediction of residues being in physical contact across interaction interfaces. Statistical inference methods detecting residue−residue coevolution have recently triggered considerable progress in using sequence data for quaternary protein structure prediction; they require, however, large joint alignments of homologous protein pairs known to interact. The generation of such alignments is a complex computational task on its own; application of coevolutionary modeling has, in turn, been restricted to proteins without paralogs, or to bacterial systems with the corresponding coding genes being colocalized in operons. Here we show that the direct coupling analysis of residue coevolution can be extended to connect the different scales, and simultaneously to match interacting paralogs, to identify interprotein residue−residue contacts and to discriminate interacting from noninteracting families in a multiprotein system. Our results extend the potential applications of coevolutionary analysis far beyond cases treatable so far.A lmost all biological processes depend on interacting proteins.Understanding protein−protein interactions is therefore key to our understanding of complex biological systems. In this context, at least two questions are of interest: First, the question "who with whom," i.e., which proteins interact; this concerns the networks connecting specific proteins inside one organism, but alsoin the context of this article-the evolutionary perspective of protein−protein interactions, which are conserved across different species. Their coevolution is at the basis of many modern computational techniques for characterizing protein−protein interactions. The second question is the question "how" proteins interact with each other, in particular, which residues are involved in the interaction interfaces, and which residues are in contact across the interfaces. Such knowledge may provide important mechanistic insight into questions related to interaction specificity or competitive interaction with partially shared interfaces.The experimental identification of protein−protein interactions is an arduous task (for reviews, cf. refs. 1 and 2): High-throughput techniques that aim to identify protein−protein interactions in vivo or in vitro are well documented and include large-scale yeast two-hybrid assays and protein affinity mass spectrometry assays. Such large-scale efforts have revealed useful information but are hampered by high false positive and false negative error rates. Structural approaches based on protein cocrystalli...

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“…Al, 2000). The crystal structures of tryptophan synthase complexed with indole-3-acetylglycine and indole-3-acetyl-l-aspartic acid revealed that both ligands bind to the active site such that the carboxylate moiety is positioned similarly as the phosphate group of the natural substrates (Weyand et al, 2002).…”

Section: Research For Finding New Target Sitesmentioning

confidence: 99%

Molecular Mechanism of Action of Herbicides

Jablonkai¹

2011

Herbicides - Mechanisms and Mode of Action

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Crystal Structures of a New Class of Allosteric Effectors Complexed to Tryptophan Synthase

Cited by 44 publications

References 43 publications

Crystal Structure of the βSer178 → Pro Mutant of Tryptophan Synthase

Crystal Structure of the βSer178 → Pro Mutant of Tryptophan Synthase

Simultaneous identification of specifically interacting paralogs and interprotein contacts by direct coupling analysis

Molecular Mechanism of Action of Herbicides

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