Thermozymes

Vieille, Claire; Burdette, Doug S.; Zeikus, J. G.

doi:10.1016/s1387-2656(08)70006-1

Cited by 157 publications

(121 citation statements)

References 312 publications

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“…The TtADPRase structure of Gd 3ϩ ternary complex was quite similar to those of the EcADPRase and MtADPRase (10,11), although the detailed structures differed. N-terminal domain ␤-sheet was shorter, loop L4 was moved outward, and loop L5 tip was situated upward by 4 Å. TtADPRase has other properties that might contribute to stability, including a lower content of chemically unstable amino acids such as Asn, Cys, and Met (25), and a smaller surface area in the N-terminal domain than for EcADPRase and MtADPRase.…”

Section: Overall Structurementioning

confidence: 98%

Structural Insights into the Thermus thermophilus ADP-ribose Pyrophosphatase Mechanism via Crystal Structures with the Bound Substrate and Metal

Yoshiba

Ooga

Nakagawa

et al. 2004

Journal of Biological Chemistry

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ADP-ribose pyrophosphatase (ADPRase) catalyzes the divalent metal ion-dependent hydrolysis of ADP-ribose to ribose 5 -phosphate and AMP. This enzyme plays a key role in regulating the intracellular ADP-ribose levels, and prevents nonenzymatic ADP-ribosylation. To elucidate the pyrophosphatase hydrolysis mechanism employed by this enzyme, structural changes occurring on binding of substrate, metal and product were investigated using crystal structures of ADPRase from an extreme thermophile, Thermus thermophilus HB8. Seven structures were determined, including that of the free enzyme, the Zn 2؉ -bound enzyme, the binary complex with ADP-ribose, the ternary complexes with ADPribose and Zn 2؉ or Gd 3؉ , and the product complexes with AMP and Mg 2؉ or with ribose 5 -phosphate and Zn 2؉ . The structural and functional studies suggested that the ADP-ribose hydrolysis pathway consists of four reaction states: bound with metal (I), metal and substrate (II), metal and substrate in the transition state (III), and products (IV). In reaction state II, Glu-82 and Glu-70 abstract a proton from a water molecule. This water molecule is situated at an ideal position to carry out nucleophilic attack on the adenosyl phosphate, as it is 3.6 Å away from the target phosphorus and almost in line with the scissile bond.Nudix pyrophosphatases are widely distributed in nature and share a highly conserved amino acid sequence motif called the "Nudix motif " (GX 5 EX 7 REUXEEXGU, where U is one of the bulky hydrophobic amino acids I, L, or V), which adopts a unique loop-helix-loop structure (1). Enzymes in this family catalyze the hydrolysis of nucleoside diphosphates, linked to another moiety x. Their postulated role is to control the cellular concentration of toxic nucleoside diphosphate derivatives or physiological metabolites, accumulation of which could be harmful (1). ADP-ribose (ADPR) 1 is one such diphosphate derivative, which is produced enzymatically as part of the turnover of NAD ϩ , cyclic ADPR, ADP-ribosylated proteins, and poly-ADP-ribosylated proteins. Although certain proteins are posttranslationally modified by ADPR, high intracellular levels of ADPR could result in nonenzymatic ADP-ribosylation. This is a deleterious process that inactivates enzymes and could interfere with the recognition of enzymatic ADP-ribosylation (2). ADPR pyrophosphatases (ADPRases) catalyze the hydrolysis of ADPR to AMP and ribose 5Ј-phosphate to prevent ADPR accumulation.ADPRase activity has been detected in all three kingdoms (3-7), but the specificity for ADPR over other substrates and the selectivity of metal ions required for activity vary between species. The mechanism underlying the different substrate specificity and the metal dependence is unknown at both the structural and functional levels. Elucidation of these properties requires the study of ADPRases from numerous sources.In this article, we investigated the catalytic mechanism of ADPRase from an extreme thermophile, Thermus thermophilus HB8 (TtADPRase). In general, proteins isolated fr...

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Section: Overall Structurementioning

confidence: 98%

Structural Insights into the Thermus thermophilus ADP-ribose Pyrophosphatase Mechanism via Crystal Structures with the Bound Substrate and Metal

Yoshiba

Ooga

Nakagawa

et al. 2004

Journal of Biological Chemistry

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“…The TPP from the mesophilic M. smegmatis, on the other hand, was completely inactivated after incubation for 0.5 min at 100°C (Klutts et al 2003), and a mixture of TPS/TPP from E. coli retained only 3% of the original activity after incubation at 50°C for 30 min (Seo et al 2000). The enhanced stability of enzymes at high temperature is the result of differences in specific amino acid composition (Vieille et al 1996). The T. thermophilus RQ-1 TPS and TPP (GenPept accession numbers AAQ16095/16096) had lower frequency of glutamine (1.4-and 4.0-fold, respectively) than their counterparts did in E. coli.…”

mentioning

confidence: 99%

Trehalose biosynthesis in Thermus thermophilus RQ-1: biochemical properties of the trehalose-6-phosphate synthase and trehalose-6-phosphate phosphatase

2004

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The genes for trehalose synthesis in Thermus thermophilus RQ-1, namely otsA [trehalose-phosphate synthase (TPS)], otsB [trehalose-phosphate phosphatase (TPP)], and treS [trehalose synthase (maltose converting) (TreS)] genes are structurally linked. The TPS/TPP pathway plays a role in osmoadaptation, since mutants unable to synthesize trehalose via this pathway were less osmotolerant, in trehalose-deprived medium, than the wild-type strain. The otsA and otsB genes have now been individually cloned and overexpressed in Escherichia coli and the corresponding recombinant enzymes purified. The apparent molecular masses of TPS and TPP were 52 and 26 kDa, respectively. The recombinant TPS utilized UDP-glucose, TDP-glucose, ADP-glucose, or GDPglucose, in this order as glucosyl donors, and glucose-6-phosphate as the glucosyl acceptor to produce trehalose-6-phosphate (T6P). The recombinant TPP catalyzed the dephosphorylation of T6P to trehalose. This enzyme also dephosphorylated G6P, and this activity was enhanced by NDP-glucose. TPS had an optimal activity at about 98°C and pH near 6.0; TPP had a maximal activity near 70°C and at pH 7.0. The enzymes were extremely thermostable: at 100°C, TPS had a half-life of 31 min, and TPP had a half-life of 40 min. The enzymes did not require the presence of divalent cations for activity; however, the presence of Co 2+ and Mg 2+stimulates both TPS and TPP. This is the first report of the characterization of TPS and TPP from a thermophilic organism.

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“…The scope of Industrial enzymes is growing because they (i) offer less polluting processes than chemical catalyst (ii) perform reactions with higher specificity than chemical catalyst and (iii) perform reaction for which chemical catalyst are not known (Claivielle et al 1996). Among these enzymes, protease is the most important enzyme and accounts for about 60% of the total worldwide sale (Gupta et al 2002).…”

Section: Introductionmentioning

confidence: 99%

Isolation and Characterization of Thermostable Protease Producing Bacteria from Soap Industry Effluent.

Thakur¹

2013

IOSR-JPBS

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Thermozymes

Cited by 157 publications

References 312 publications

Structural Insights into the Thermus thermophilus ADP-ribose Pyrophosphatase Mechanism via Crystal Structures with the Bound Substrate and Metal

Structural Insights into the Thermus thermophilus ADP-ribose Pyrophosphatase Mechanism via Crystal Structures with the Bound Substrate and Metal

Trehalose biosynthesis in Thermus thermophilus RQ-1: biochemical properties of the trehalose-6-phosphate synthase and trehalose-6-phosphate phosphatase

Isolation and Characterization of Thermostable Protease Producing Bacteria from Soap Industry Effluent.

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