Mechanism-Based Inhibitors of Cytokinin Oxidase/Dehydrogenase Attack FAD Cofactor

Analysis of N-glycosylation in maize cytokinin oxidase/dehydrogenase 1 using a manual microgradient chromatographic separation coupled offline to MALDI-TOF/TOF mass spectrometry

“…Both peaks were accompanied by neighbors without oxidized methionine (m/z 4855 and 4316, respectively). The other N-glycopeptides with m/z 5136 and 5740 (average masses) then refer to the peptides 323-367 and 49-101 (including an amino acid substitution of Gly79 to Ala79 [36]), each with two remaining GlcNAc residues at their respective N-glycosylation sites. MALDI-TOF/TOF MS measurements revealed monoisotopic patterns of the mentioned peptide peaks with signal-to-noise ratios above 20 and mass accuracy less than 100 ppm with external calibration (not shown).…”

The Biotechnological Potential of Cytokinin Status Manipulation

“…In the most cases, their modes of binding to CKX have been clarified by structural analysis (Kopečný et al 2010). The activity of CKX was shown to be strongly inhibited by unsaturated allenic (HA-1, HA-8) and alkyne substrate analogues (Houba-Hérin et al 1999;Suttle and Mornet 2005;Kopečný et al 2008). Kinetic and X-ray crystallographic studies proved that HA-1 and HA-8 behave as mechanism-based inhibitors, and it was therefore suggested that they might be useful tools for regulating CKX activity in plants (Suttle and Mornet 2005;Kopečný et al 2008).…”

“…The activity of CKX was shown to be strongly inhibited by unsaturated allenic (HA-1, HA-8) and alkyne substrate analogues (Houba-Hérin et al 1999;Suttle and Mornet 2005;Kopečný et al 2008). Kinetic and X-ray crystallographic studies proved that HA-1 and HA-8 behave as mechanism-based inhibitors, and it was therefore suggested that they might be useful tools for regulating CKX activity in plants (Suttle and Mornet 2005;Kopečný et al 2008). A class of substituted 6-anilinopurines was recently identified as new and potent CKX inhibitors, and the molecular basis of their interaction with CKX was described (Zatloukal et al 2008;Spíchal et al unpublished results).…”

Cytokinin activity of disubstituted aminopurines in Amaranthus

“…An effect on CK metabolism could explain their activity at concentrations lower than 50 mM. Recently, Kopečný et al (2008) reported several CK analogues with the ability to inactivate CKO, and that these compounds behaved as ''suicide substrates'', as they irreversibly bind to the enzyme cofactor FAD. However, the possibility for benzyl-alkyl adenines to act in a similar way is quite remote, since their chemical structures do not permit the in situ formation of a Michael system by an oxidative mechanism, which is necessary for an irreversible bind of the CK moiety to oxidases.…”