Crystal Structures of a Mutant (βK87T) Tryptophan Synthase α<sub>2</sub>β<sub>2</sub> Complex with Ligands Bound to the Active Sites of the α- and β-Subunits Reveal Ligand-Induced Conformational Changes

Rhee, Sangkee; Parris, Kevin; Hyde, C. Craig; Ahmed, Salman; Miles, Edith Wilson; Davies, David R.

doi:10.1021/bi9700429

Cited by 155 publications

(291 citation statements)

References 48 publications

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“…In Figure 8, we show in pink the region exhibiting this pronounced hindered mobility in the presence of substrates, and in magenta its most strongly affected segments ( 130-145 and 155-170). This region ( 103-187) closely matches the so-called COMM domain of the -subunit which was pointed out by Schneider et al (7), as a slight modification of the "mobile region" originally pointed out by Rhee et al (5), to play an important role in the allosteric communication between the R-and -sites. The most severely affected regions identified here ( 130-145 and 155-170) will be referred to as the COMM core regions below.…”

Section: Changes In Mobilities Induced Upon Ligand Bindingsupporting

confidence: 71%

“…This is the only currently available structure in which residues participating in loop RL6 are partly visible. This feature was attributed to the presence of substrates bound on both subunits (5). The obstruction in conformational mobility of RL6 upon binding of ligands was also indicated by the prevention of tryptic cleavage (41).…”

Section: Discussionmentioning

confidence: 96%

“…Results for only subunits R 1 and 1 are displayed; those for subunits R 2 and 2 are identical. Experimental data were taken from PDB file 2tys deposited by Rhee et al (5). Theoretical results were calculated using eqs 2 and 4.…”

Section: Global Motion Driven By the First Principal Mode Of Actionmentioning

confidence: 99%

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Cooperative Fluctuations and Subunit Communication in Tryptophan Synthase

Bahar

Jernigan

1999

Biochemistry

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Tryptophan synthase (TRPS), with linearly arrayed subunits R R, catalyzes the last two reactions in the biosynthesis of L-tryptophan. The two reactions take place in the respective R-and -subunits of the enzyme, and the intermediate product, indole, is transferred from the R-to the -site through a 25 Å long hydrophobic tunnel. The occurrence of a unique ligand-mediated long-range cooperativity for substrate channeling, and a quest to understand the mechanism of allosteric control and coordination in metabolic cycles, have motivated many experimental studies on the structure and catalytic activity of the TRPS R 2 2 complex and its mutants. The dynamics of these complexes are analyzed here using a simple but rigorous theoretical approach, the Gaussian network model. Both wild-type and mutant structures, in the unliganded and various liganded forms, are considered. The substrate binding site in the -subunit is found to be closely coupled to a group of hinge residues ( 77-89 and 376-379) near the -interface. These residues simultaneously control the anticorrelated motion of the two -subunits, and the opening or closing of the hydrophobic tunnel. The latter process is achieved by the large amplitude fluctuations of the so-called COMM domain in the same subunit. Intersubunit communications are strengthened in the presence of external aldimines bound to the -site. The motions of the COMM core residues are coordinated with those of the R-hinge residues 174-179 on the interfacial helix H6 at the entrance of the hydrophobic tunnel. And the motions of H6 are coupled, via helix H1 and RL6, to those of the loop RL2 that includes the R-subunit catalytically active residue Asp60. Overall, our analysis sheds light on the molecular machinery underlying subunit communication, and identifies the residues playing a key role in the cooperative transmission of conformational motions across the two reaction sites.The tryptophan synthase (TRPS) 1 R 2 2 complex is a bifunctional enzyme that catalyzes the last two reactions in the biosynthesis of L-tryptophan (L-Trp). The bacterial enzyme structure consists of two R-and two -subunits arranged in an extended R R order (Figure 1a,b). The Rand -subunits contain the respective sites for the R-and -reactions producing L-Trp: the cleavage of indole 3-glycerol phosphate (IGP) to release indole and glyceraldehyde 3-phosphate (G3P) (R-reaction) and the conversion of indole to L-Trp via a condensation reaction with L-serine ( -reaction). The latter is mediated by the cofactor pyridoxal 5′-phosphate (PLP), which forms with L-Ser a quasi-stable R-aminoacrylate intermediate, E(A-A), highly reactive toward indole. The R -reaction resulting from the combination of presents a typical example of long-range allosteric effect in that the two reactions, occurring at two topologically distinct and distant sites, are highly coordinated through a PLPdependent activation-deactivation mechanism, and the metabolite, indole, is transferred from the active site in the R-subunit to the other in the -subuni...

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Section: Changes In Mobilities Induced Upon Ligand Bindingsupporting

confidence: 71%

Section: Discussionmentioning

confidence: 96%

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Cooperative Fluctuations and Subunit Communication in Tryptophan Synthase

Bahar

Jernigan

1999

Biochemistry

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“…Present investigation has been correlated with the many studies made by various scientists, [17][18][19][20][21][22][23][24][25][26][27][28]. The studies supports the findings of the present study specially the sequence and structure analyzed against the standards taken (Pfu beta 2 subunit with Stm beta subunit of tryptophan synthase) for sequence analysis and structure prediction.…”

Section: Discussionsupporting

confidence: 88%

Comparative Study of Tryptophan Synthase Beta Chainbetween Pyrococcus furiosus (Archaea) and Salmonella typhimurium (Proteobacteria)

Bisht¹,

Behera²,

Panda³

2011

IJBBB

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Abstract-The present investigation deals with comparative study of beta chain of the enzyme tryptophan synthase from Salmonella typhimurium and Pyrococcus furiosus. The study yielded few significant results that the sequence of beta 2 subunit (PF1706) from Pfu is closely related to the beta chain of Salmonella typhimurium in comparison to the beta 1. The phylogenetics of these two strains indicates that the beta 2 subunit plays a vital role in tryptophan synthesis in Pyrococcus furiosus. In the present investigation Salmonella typhimurium was used as a reference organism for comparative genomics of tryptophan synthase beta chain amongst a group of organisms 13 archaea and 41 proteobacteria respectively. The study reveals that most of the sequences of archaea are distantly related and was observed that there are certain sequences like pfu PF1706 which are closer to Stm that is a proteobacteria.

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“…Catalytic rates of ϳ135 s Ϫ1 (29,30) are consistent with turnover-limiting protein conformational changes (20,(31)(32)(33). Paired substratefree and transition state analog complex (TSAC) structures are available for both arginine kinase and creatine kinase; the TSAC has bound phosphagen, ADP, and a nitrate mimicking the ␥-phosphoryl in transit (21,35).…”

mentioning

confidence: 80%

The Structure of Lombricine Kinase

Bush

Kirillova

Clark

et al. 2011

Journal of Biological Chemistry

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Lombricine kinase is a member of the phosphagen kinase family and a homolog of creatine and arginine kinases, enzymes responsible for buffering cellular ATP levels. Structures of lombricine kinase from the marine worm Urechis caupo were determined by x-ray crystallography. One form was crystallized as a nucleotide complex, and the other was substrate-free. The two structures are similar to each other and more similar to the substrate-free forms of homologs than to the substrate-bound forms of the other phosphagen kinases. Active site specificity loop 309 -317, which is disordered in substrate-free structures of homologs and is known from the NMR of arginine kinase to be inherently dynamic, is resolved in both lombricine kinase structures, providing an improved basis for understanding the loop dynamics. Phosphagen kinases undergo a segmented closing on substrate binding, but the lombricine kinase ADP complex is in the open form more typical of substrate-free homologs. Through a comparison with prior complexes of intermediate structure, a correlation was revealed between the overall enzyme conformation and the substrate interactions of His 178 . Comparative modeling provides a rationale for the more relaxed specificity of these kinases, of which the natural substrates are among the largest of the phosphagen substrates.Lombricine, arginine, and creatine kinases (EC 2.7.3) are homologous phosphagen kinases that catalyze the buffering of cellular ATP levels through phosphoryl transfer to/from their respective guanidino-containing substrates. The reaction is central to short-term temporal energy buffering (1, 2) as well as in spatial shuttling of energy from production to consumption sites (3-5). A wide array of endergonic processes is driven by nucleotide hydrolysis, from motion in molecular motors, active transport, and synthetic metabolism to signal transduction. Thus, the maintenance of a constant ATP/ADP ratio, displaced far from thermodynamic equilibrium in the face of high and variable rates of ATP turnover, is crucial for cellular homeostasis (2).Different organisms use different phosphagen substrates, usually only one and each with its own specific phosphagen kinase ( Fig. 1) (2). Lombricine kinase, as well as taurocyamine kinase and glycocyamine kinase, is found exclusively in annelids and allied groups (2). Phylogenetic analyses and studies of the intron/exon organization of the genes of these phosphagen kinases unique to annelids have shown that they are more closely related to creatine kinases (with which they share 50 -60% sequence identity) than typical monomeric arginine kinases such as that from the horseshoe crab (6) (40% sequence identity). Annelids are more diverse in their choice of phosphagen, and the substrate specificities of the corresponding kinases are often lower (6).Structural work has concentrated on the presumptive ancestral arginine kinase and the vertebrate creatine kinase (7, 8), but sequence alignment suggests that subunit fold, if not quaternary structure, is conserved across the fa...

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Crystal Structures of a Mutant (βK87T) Tryptophan Synthase α₂β₂ Complex with Ligands Bound to the Active Sites of the α- and β-Subunits Reveal Ligand-Induced Conformational Changes

Cited by 155 publications

References 48 publications

Cooperative Fluctuations and Subunit Communication in Tryptophan Synthase

Cooperative Fluctuations and Subunit Communication in Tryptophan Synthase

Comparative Study of Tryptophan Synthase Beta Chainbetween Pyrococcus furiosus (Archaea) and Salmonella typhimurium (Proteobacteria)

The Structure of Lombricine Kinase

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Crystal Structures of a Mutant (βK87T) Tryptophan Synthase α2β2 Complex with Ligands Bound to the Active Sites of the α- and β-Subunits Reveal Ligand-Induced Conformational Changes

Cited by 155 publications

References 48 publications

Cooperative Fluctuations and Subunit Communication in Tryptophan Synthase

Cooperative Fluctuations and Subunit Communication in Tryptophan Synthase

Comparative Study of Tryptophan Synthase Beta Chainbetween Pyrococcus furiosus (Archaea) and Salmonella typhimurium (Proteobacteria)

The Structure of Lombricine Kinase

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Product

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Crystal Structures of a Mutant (βK87T) Tryptophan Synthase α₂β₂ Complex with Ligands Bound to the Active Sites of the α- and β-Subunits Reveal Ligand-Induced Conformational Changes