Kinetic and spectroscopic studies of the ATP:corrinoid adenosyltransferase PduO from Lactobacillus reuteri: substrate specificity and insights into the mechanism of Co(II)corrinoid reduction

Biochemistry. 2008 Aug 26;47(34):9007-15. doi: 10.1021/bi800419e. Epub 2008 Aug 2.


The PduO-type ATP:corrinoid adenosyltransferase from Lactobacillus reuteri ( LrPduO) catalyzes the formation of the essential Co-C bond of adenosylcobalamin (coenzyme B 12) by transferring the adenosyl group from cosubstrate ATP to a transient Co (1+)corrinoid species generated in the enzyme active site. While PduO-type enzymes have previously been believed to be capable of adenosylating only Co (1+)cobalamin (Co (1+)Cbl (-)), our kinetic data obtained in this study provide in vitro evidence that LrPduO can in fact also utilize the incomplete corrinoid Co (1+)cobinamide (Co (1+)Cbi) as an alternative substrate. To explore the mechanism by which LrPduO overcomes the thermodynamically challenging reduction of its Co (2+)corrinoid substrates, we have examined how the enzyme active site alters the geometric and electronic properties of Co (2+)Cbl and Co (2+)Cbi (+) by using electronic absorption, magnetic circular dichroism, and electron paramagnetic resonance spectroscopic techniques. Our data reveal that upon binding to LrPduO that was preincubated with ATP, both Co (2+)corrinoids undergo a partial ( approximately 40-50%) conversion to distinct paramagnetic Co (2+) species. The spectroscopic signatures of these species are consistent with essentially four-coordinate, square-planar Co (2+) complexes, based on a comparison with the results obtained in our previous studies of related enzymes. Consequently, it appears that the general strategy employed by adenosyltransferases for effecting Co (2+) --> Co (1+) reduction involves the formation of an "activated" Co (2+)corrinoid intermediate that lacks any significant axial bonding interactions, to stabilize the redox-active, Co 3d z (2) -based molecular orbital.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Adenosine Triphosphate / metabolism
  • Alkyl and Aryl Transferases / chemistry*
  • Alkyl and Aryl Transferases / metabolism
  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / metabolism
  • Binding Sites
  • Cobalt / chemistry*
  • Cobalt / metabolism
  • Cobamides / chemistry
  • Cobamides / metabolism
  • Corrinoids / chemistry*
  • Corrinoids / metabolism
  • Kinetics
  • Limosilactobacillus reuteri / enzymology*
  • Oxidation-Reduction
  • Substrate Specificity
  • Thermodynamics


  • Bacterial Proteins
  • Cobamides
  • Corrinoids
  • Cobalt
  • Adenosine Triphosphate
  • Alkyl and Aryl Transferases
  • ATP-corrinoid adenosyltransferase
  • cobamamide