Catalytic mechanism of xylose (glucose) isomerase from Clostridium thermosulfurogenes. Characterization of the structural gene and function of active site histidine.

Lee, C Y; Bagdasarian, Michael; Meng, M.; Zeikus, J. G.

doi:10.1016/s0021-9258(17)30628-2

Cited by 87 publications

(28 citation statements)

References 38 publications

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“…Fig. 3(A) shows that, at pH levels below 6.0 and above 7.5, V max of TnapXI dropped sharply at acidic and basic limbs because, after receiving H + , the capacity of imidazole nitrogen for the formation of hydrogen bonding with the substrate was reduced; this is in accordance with results previously calculated for thermophilic xylose isomerase from Thermoanaerobacterium thermosulfurogenes (TTXI) (formerly known as Clostridium thermosulfurogenes 4B) 47 . The E a of TnapXI is in close agreement with the 80 kJ.mol −1 reported for TNXI.…”

Section: Discussionsupporting

confidence: 86%

Kinetics and thermodynamics for aldo/keto conversion of D‐xylose and D‐glucose catalyzed by xylose isomerase from Thermotoga naphthophilaRKU‐10

Fatima

Aftab

Haq

2021

J of Chemical Tech & Biotech

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BACKGROUND Aldo‐keto isomerizations of D‐xylose and D‐glucose catalyzed by xylose isomerase (XI) is commercially carried out to produce bio‐ethanol and high fructose corn syrup. XI is a metalloenzyme. In this paper, kinetics and thermodynamics of the activity and stability of hyperthermophilic recombinant XI from Thermotoga naphthophila RKU‐10T (TnapXI) were studied in the presence and absence of metal ions. RESULTS Kinetics parameters of recombinant XI from TnapXI were calculated as Km = 0.99 mmol L–1, Vmax = 500 μmol.mg−1.min−1, and kcat = 68.3 min−1 for the isomerization of D‐xylose(aldo)⇌D‐xylulose(keto), whereas for D‐glucose(aldo)⇌D‐fructose(keto) conversion Km = 7.72 mmol L–1, Vmax = 90 μmol.mg−1.min−1, and kcat = 12.4 min−1. Ionization constants were calculated as pKa1 = 6.0 and pKa2 = 7.6, whereas activation constants (Ka) of TnapXI were 0.3, 0.5, and 4.0 mmol L–1 for Co2+, Mn2+, and Mg2+, respectively. Inhibition constant (Ki) was 0.119 mmol L–1 for Ca2+. Q10 was 2.8 and Ea = 82.25 kJ.mol−1. Thermodynamics for isomerization were calculated as ∆H* = 79.2 kJ.mol−1, ∆G* = 83 kJ.mol−1, and ∆S* = −10.5 J.mol−1.K−1. The z‐value was 12.6 °C for apo‐TnapXI and 32.6 °C for holo‐TnapXI. Deactivation parameters were ∆G*(d) = 100 kJ.mol−1, ∆H*(d) = 206 kJ.mol−1, ∆S*(d) = 0.28 kJ.mol−1.K−1 for apo‐TnapXI and t1/2 = 18 min at 95 °C, while ∆G*(d) = 104 kJ.mol−1, ∆H*(d) = 767 kJ.mol−1, ∆S*(d) = 1.8 kJ.mol−1.K−1 for holo‐TnapXI and t1/2 = 65 min at 95 °C. CONCLUSION These noteworthy features make this enzyme an attractive candidate for the industrial production of fuel ethanol and high fructose corn syrup at high temperatures to maximize the product yield. © 2021 Society of Chemical Industry (SCI).

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Section: Discussionsupporting

confidence: 86%

Kinetics and thermodynamics for aldo/keto conversion of D‐xylose and D‐glucose catalyzed by xylose isomerase from Thermotoga naphthophilaRKU‐10

Fatima

Aftab

Haq

2021

J of Chemical Tech & Biotech

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“…The kinetic isotope effect was calculated to be 1.80 without tunneling and 3.75 with tunneling. The latter is within the range expected from various experimental [89] determinations.…”

Section: Xylose Isomerasesupporting

confidence: 87%

Variational Transition State Theory in the Treatment of Hydrogen Transfer Reactions

Truhlar¹,

Garrett²

2006

Hydrogen‐Transfer Reactions

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“…The incorporation of solvent protons (tritium) into the fructose product of D-xylose isomerase has been reported as <0.4% (Rose et al, 1969). Although such a low exchange rate has led to the suggestion that the mechanism of D-xylose isomerase proceeds via a hydride shift (Farber et al, 1989;Lee et al, 1990;Whitlow etal., 1991) it has been pointed out for the enzyme glyoxylase I that such results do not rule out a fast proton transfer taking place in a shielded active site (Hall et al, 1976). However, this possibility can be tested by observing the exchange at increased temperatures (Rose & O'Connell, 1961).…”

Section: Resultsmentioning

confidence: 99%

“…Our suggestion was based in part on the observation that the substrate was coordinated directly to one of the two metal sites, which suggested a possible analogy with metal-promoted hydride transfer reactions such as the Meerwein-Ponndorf-Verley-Oppenheimer reaction (Kemp & Vellaccio, 1980). Subsequently, high-resolution crystal structures of enzyme-substrate and enzyme-inhibitor complexes for D-xylose isomerases from various organisms have been interpreted in terms of a hydridetransfer mechanism and detailed roles for the metals and active site residues have been proposed Lee et al, 1990;Whitlow etal., 1991;Jenkins etal., 1992;Rangarajan & Hartley, 1992;. All of these structural data are indeed inconsistent with a proton-transfer mechanism [with one exception (Carrel etal., 1989); however, this interpretation has been challenged ].…”

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confidence: 99%

Isotopic Exchange plus Substrate and Inhibition Kinetics of D-Xylose Isomerase Do Not Support a Proton-Transfer Mechanism

et al. 1994

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The D-xylose isomerase of Streptomyces olivochromogenes is a Mg2+- or Mn(2+)-dependent enzyme that catalyzes the aldose-ketose isomerization of xylose to xylulose or of glucose to fructose. Proton exchange into water during enzyme-catalyzed isomerization of C-2 tritiated glucose at 15, 25 and 55 degrees C shows < 0.6% exchange (the loss of one proton in every billion turnovers). High concentrations of guanidine hydrochloride and extremes of pH had no effect on the amount of exchange detected. Such a low percentage of exchange is inconsistent with a proton-transfer mechanism as the main kinetic pathway for isomerization. 19F NMR experiments showed no release of fluoride after incubation of the enzyme for 4 weeks with 800 mM 3-deoxy-3-fluoroglucose or 3-deoxy-3-fluoroallose (both are competitive inhibitors with Ki values of 600 mM). This result is also inconsistent with a proton-transfer mechanism. A hydride-shift mechanism following ring opening has been proposed for the isomerization. Enzyme-catalyzed ring opening was directly measured by demonstrating H2S release upon reaction of xylose isomerase with 1-thioglucose. D-Xylose isomerase-catalyzed interconversion of glucose to fructose exhibited linear Arrhenius behavior with an activation energy of 14 kcal/mol from 0 to 50 degrees C. No change in rate-determining step occurs over this temperature range. 13C NMR experiments with glucose show that enzyme-bound magnesium or manganese does not interact specifically with any one site on the sugar. These results are consistent with nonproductive binding modes for the substrate glucose in addition to productive binding.

show abstract

Catalytic mechanism of xylose (glucose) isomerase from Clostridium thermosulfurogenes. Characterization of the structural gene and function of active site histidine.

Cited by 87 publications

References 38 publications

Kinetics and thermodynamics for aldo/keto conversion of D‐xylose and D‐glucose catalyzed by xylose isomerase from Thermotoga naphthophilaRKU‐10

Kinetics and thermodynamics for aldo/keto conversion of D‐xylose and D‐glucose catalyzed by xylose isomerase from Thermotoga naphthophilaRKU‐10

Variational Transition State Theory in the Treatment of Hydrogen Transfer Reactions

Isotopic Exchange plus Substrate and Inhibition Kinetics of D-Xylose Isomerase Do Not Support a Proton-Transfer Mechanism

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