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Volumn 125, Issue 36, 2003, Pages 10877-10884

The first experimental test of the hypothesis that enzymes have evolved to enhance hydrogen tunneling

Author keywords

[No Author keywords available]

Indexed keywords

QUANTUM MECHANICAL TUNNELING;

EID: 0043236031     PISSN: 00027863     EISSN: None     Source Type: Journal    
DOI: 10.1021/ja030120h     Document Type: Article
Times cited : (92)

References (90)
  • 11
    • 0042907100 scopus 로고    scopus 로고
    • note
    • (a) An early version of this hypothesis dates back to a 1971 report by Gold, who studied "facilitated proton transfers" in chymotrypsin and carbonic anhydrase. Gold notes, "...enzymes may act by enforcing small reductions in the length of hydrogen bonds, thereby increasing the probability of quantum mechanical tunneling."
  • 12
    • 0003960770 scopus 로고
    • John Wiley and Sons: New York
    • (b) Moore and Pearson have noted, "It is correct to say that tunneling is always a factor in reactions involving transfer of hydrogen atoms, protons, or hydride ions." Moore, J. W.; Pearson, R. G. Kinetics and Mechanism, 3rd ed.; John Wiley and Sons: New York, 1981; p. 371. The extent of tunneling might, of course, be nearly complete at low temperatures and for penetrable barriers to very little at higher temperatures for barriers that preclude significant tunneling.
    • (1981) Kinetics and Mechanism, 3rd Ed. , pp. 371
    • Moore, J.W.1    Pearson, R.G.2
  • 13
    • 0014431004 scopus 로고
    • Wang, J. H. Science 1968, 161, 328.
    • (1968) Science , vol.161 , pp. 328
    • Wang, J.H.1
  • 30
    • 0041905042 scopus 로고    scopus 로고
    • note
    • It has been noted by Bruice and Benkovic, "There is at present, however, little direct evidence as to how enzymes use correlated motion in catalysis. Theoretical treatments have been offered which explicitly recognize the role of protein dynamics in the treatment of hydride transfer reactions and tunneling."
  • 31
  • 43
    • 0042406295 scopus 로고    scopus 로고
    • note
    • The reason for the ratio of less than one in tunneling cases can be seen by coplotting Arrhenius plots of In k vs 1/T for the hydrogen and deuterium cases. If the tunneling component is large, the rate will be enhanced at low temperatures, causing significant curvature in the hydrogen case. This will make the pre-exponential factor (calculated from the intercept of this line) lower, and the ratio of pre-exponential factors less than one.
  • 44
    • 0042907089 scopus 로고    scopus 로고
    • note
    • There is evidence that indicates that the Co-C bond cleavage and the hydrogen atom abstraction are coupled. Note, however, that our data showing identical tunneling criteria with and without the enzyme indicate substantial adenosyl radical character in the hydrogen atom abstraction reaction.
  • 47
    • 0042406293 scopus 로고    scopus 로고
    • note
    • There is some nondefinitive data for QM tunneling in diol dehydratase, ethanolamine, ammonia lyase, and glutamate mutase. See sections S-3 through S-5 of the Supporting Information for further discussion.
  • 50
    • 0042406291 scopus 로고    scopus 로고
    • note
    • A small amount of the adenine (∼5%) is also observed due to a well-established, competitive, pH-dependent Co-C heterolysis.
  • 51
    • 0041403930 scopus 로고    scopus 로고
    • note
    • The most probable mechanism for the H atom loss reaction after the cyclization step involves H atom abstraction from the cyclized nucleoside radical by the persistent cobalamin radical. The possible disproportionation of the cyclized nucleoside radical is probably never kinetically viable once even a small amount of Co(II)Cbl builds Up.
  • 54
    • 0041905031 scopus 로고    scopus 로고
    • note
    • 6 (section S-1 in the Supporting Information).
  • 59
    • 0042406288 scopus 로고    scopus 로고
    • note
    • The required extrapolation of our In KIE vs 1/T plot to lower temperatures rather than measured experimentally leads to an estimated lower limit of the KIE of 29 at 20 °C. similar to the value of 35.6 reported for methylmalonyl-CoA mutase. Alternatively, an extrapolation of the enzymic data to higher temperatures leads to an estimated enzymic KIE of 12.7 at 80 °C, again within experimental error of our enzyme-free KIE of 12.4 ± 1.1. We have also examined a system, β-neopentylcobalamin, where thermolysis occurs in the same temperature range in which the enzymic tunneling data was obtained. That system, again, shows no difference within experimental error for the three tunneling criteria vs those for methylmalonyl-CoA mutase.
  • 82
    • 0041791031 scopus 로고    scopus 로고
    • note
    • -1).
  • 87
    • 0041290432 scopus 로고    scopus 로고
    • submitted for publication
    • +-transfer distance and increasing the tunneling mode anharmonicity, the two most important parameters in their tunneling model.
    • (2003) J. Phys. Chem.
    • Siebrand, W.1    Smedarchina, Z.2
  • 88
    • 0037123216 scopus 로고    scopus 로고
    • (b) Knapp, M. J.; Rickert, K. W.; Klinman, J. P. J. Am. Chem. Soc. 2002, 124, 3865, Siebrand and Snedarchina show that there is no statistically valid difference in the data in this paper for mutant vs wild-type enzyme (because of the too-narrow temperature range over which the data could be obtained), thereby negating the claimed support in this paper for the putative protein-squeezing mechanism.
    • (2002) J. Am. Chem. Soc. , vol.124 , pp. 3865
    • Knapp, M.J.1    Rickert, K.W.2    Klinman, J.P.3
  • 89
    • 0042292197 scopus 로고    scopus 로고
    • note
    • Two of the HPLC traces contained a small unassigned peak (retention time ∼7.7 min). It did not have a UV-visible signal at 525 nm, indicating that it is not a cobalamin. It is suspected, but not proven, that it is a nucleoside formed by the cycloadenosyl radical that has not completely converted to c-Ado. Because this peak was only present in two out of 34 samples, and since there was no corresponding peak in the deuterated ethylene glycol system, chromatograms with this peak were not used in the calculation of the ratio of c-Ado/Ado-H.
  • 90
    • 0042793170 scopus 로고    scopus 로고
    • note
    • Since it is possible that the m/z window setting on the mass spectrometer could have affected the observed product ratios by changing the number of ions captured in the ion trap, a control experiment varying the m/z window setting was performed. An 8-MeOAdoCbl thermolysis solution was analyzed by MS-HPLC with the 100-1650 m/z window and then immediately reanalyzed with the MS-HPLC set to monitor only a 270-300 m/z window. The results were the same within experimental error.


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