(R)-Roscovitine (CYC202) is often referred to as a "selective inhibitor of cyclin-dependent kinases." Besides its use as a biological tool in cell cycle, neuronal functions, and apoptosis studies, it is currently evaluated as a potential drug to treat cancers, neurodegenerative diseases, viral infections, and glomerulonephritis. We have investigated the selectivity of (R)-roscovitine using three different methods: 1) testing on a wide panel of purified kinases that, along with previously published data, now reaches 151 kinases; 2) identifying roscovitine-binding proteins from various tissue and cell types following their affinity chromatography purification on immobilized roscovitine; 3) investigating the effects of roscovitine on cells deprived of one of its targets, CDK2. Altogether, the results show that (R)-roscovitine is rather selective for CDKs, in fact most kinases are not affected. However, it binds an unexpected, non-protein kinase target, pyridoxal kinase, the enzyme responsible for phosphorylation and activation of vitamin B 6 . These results could help in interpreting the cellular actions of (R)-roscovitine but also in guiding the synthesis of more selective roscovitine analogs.

The three-dimensional structures of brain pyridoxal kinase and its complex with the nucleotide ATP have been elucidated in the dimeric form at 2.1 and 2.6 Å, respectively. Results have shown that pyridoxal kinase, as an enzyme obeying random sequential kinetics in catalysis, does not possess a lid shape structure common to all kinases in the ribokinase superfamily. This finding has been shown to be in line with the condition that pyridoxal kinase binds substrates with variable sizes of chemical groups at position 4 of vitamin B 6 and its derivatives. In addition, the enzyme contains a 12-residue peptide loop in the active site for the prevention of premature hydrolysis of ATP. Conserved amino acid residues Asp 118 and Tyr 127 in the peptide loop could be moved to a position covering the nucleotide after its binding so that its chance to hydrolyze in the aqueous environment of the active site was reduced. With respect to the evolutionary trend of kinase enzymes, the existence of this loop in pyridoxal kinase could be classified as an independent category in the ribokinase superfamily according to the structural feature found and mechanism followed in catalysis.

Pyridoxal kinase (PDXK) catalyzes the phosphorylation of pyridoxal, pyridoxamine, and pyridoxine in the presence of ATP and Zn 2؉ . This constitutes an essential step in the synthesis of pyridoxal 5-phosphate (PLP), the active form of vitamin B 6 , a cofactor for over 140 enzymes. (R)-Roscovitine (CYC202, Seliciclib) is a relatively selective inhibitor of cyclin-dependent kinases (CDKs), currently evaluated for the treatment of cancers, neurodegenerative disorders, renal diseases, and several viral infections. Affinity chromatography investigations have shown that (R)-roscovitine also interacts with PDXK. To understand this interaction, we determined the crystal structure of PDXK in complex with (R)-roscovitine, N 6 -methyl-(R)-roscovitine, and O 6 -(R)-roscovitine, the two latter derivatives being designed to bind to PDXK but not to CDKs. Structural analysis revealed that these three roscovitines bind similarly in the pyridoxal-binding site of PDXK rather than in the anticipated ATP-binding site. The pyridoxal pocket has thus an unexpected ability to accommodate molecules different from and larger than pyridoxal. This work provides detailed structural information on the interactions between PDXK and roscovitine and analogs. It could also aid in the design of roscovitine derivatives displaying strict selectivity for either PDXK or CDKs.

The crystal structure of allophycocyanin from red algae Porphyra yezoensis (APC-PY) at 2.2-Å resolution has been determined by the molecular replacement method. In the crystal, two APC-PY trimers associate face to face into a hexamer. The assembly of two trimers within the hexamer is similar to that of C-phycocyanin (C-PC) and R-phycoerythrin (R-PE) hexamers, but the assembly tightness of the two trimers to the hexamer is not so high as that in C-PC and R-PE hexamers.The chromophore-protein interactions and possible pathway of energy transfer were discussed. Phycocyanobilin 1␣84 of APC-PY forms 5 hydrogen bonds with 3 residues in subunit 2␤ of another monomer. In R-PE and C-PC, chromophore 1␣84 only forms 1 hydrogen bond with 2␤77 residue in subunit 2␤. This result may support and explain great spectrum difference exists between APC trimer and monomer.Phycobilisomes are large supramolecular aggregates attached to the stromal side of the thylakoid membrane in cyanobacteria, red algae, and cryptomonads. These supramolecular aggregates are light-harvesting protein pigment complexes that are composed of phycobiliproteins and linker proteins.