Phytochrome B integrates light and temperature signals in Arabidopsis.

Ambient temperature regulates many aspects of plant growth and development, but its sensors are unknown. Here, we demonstrate that the phytochrome B (phyB) photoreceptor participates in temperature perception through its temperature-dependent reversion from the active Pfr state to the inactive Pr state. Increased rates of thermal reversion upon exposing Arabidopsis seedlings to warm environments reduce both the abundance of the biologically active Pfr-Pfr dimer pool of phyB and the size of the associated nuclear bodies, even in daylight. Mathematical analysis of stem growth for seedlings expressing wild-type phyB or thermally stable variants under various combinations of light and temperature revealed that phyB is physiologically responsive to both signals. We therefore propose that in addition to its photoreceptor functions, phyB is a temperature sensor in plants.

Three-dimensional structure of DesVI from Streptomyces venezuelae: a sugar N,N-dimethyltransferase required for dTDP-desosamine biosynthesis.

D-Desosamine, or 3-(dimethylamino)-3,4,6-trideoxyglucose, is an unusual sugar found on the macrolide antibiotic erythromycin, and it has been shown to play a critical role in the biological activity of the drug. Desosamine is added to the parent aglycone via the action of a glycosyltransferase that utilizes dTDP-desosamine as its substrate. Six enzymes are required for the biosynthesis of dTDP-desosamine in Streptomyces venezuelae, with the last step catalyzed by DesVI, an N, N-dimethyltransferase. Here we describe the X-ray crystal structure determined to 2.0 A resolution of DesVI complexed with S-adenosylmethionine (SAM) and the substrate analogue UDP-benzene. Each subunit of the DesVI dimer contains a seven-stranded mixed beta-sheet flanked on either side by alpha-helices. In addition to this major tertiary structural element, there is a four-stranded antiparallel beta-sheet that provides the platform necessary for subunit-subunit assembly. On the basis of the UDP-benzene binding mode, the DesVI substrate, dTDP-3-(methylamino)-3,4,6-trideoxyglucose, has been modeled into the active site. This model places the C-6' methyl group of the sugar into a hydrophobic patch that is well-conserved among putative nucleotide-linked sugar dimethyltransferases. It is formed by Trp 140, Met 178, and Ile 200. The sugar C-2' hydroxyl sits near Tyr 14, and its C-3' amino group is properly positioned for direct in-line attack of the cofactor's reactive methyl group. While methyltransferases that catalyze single alkylations at carbons, oxygens, sulfurs, and nitrogens have been well characterized, little is known regarding enzymes capable of N,N-dimethylation reactions. As such, the ternary structure of DesVI reported here serves as a structural paradigm for a new family of dimethyltransferases that function on nucleotide-linked sugars.

Molecular architecture of DesV from Streptomyces venezuelae: a PLP-dependent transaminase involved in the biosynthesis of the unusual sugar desosamine.

Desosamine is a 3-(dimethylamino)-3,4,6-trideoxyhexose found in certain macrolide antibiotics such as the commonly prescribed erythromycin. Six enzymes are required for its biosynthesis in Streptomyces venezuelae. The focus of this article is DesV, which catalyzes the PLP-dependent replacement of a 3-keto group with an amino functionality in the fifth step of the pathway. For this study the three-dimensional structures of both the internal aldimine and the ketimine intermediate with glutamate were determined to 2.05 A resolution. DesV is a homodimer with each subunit containing 12 alpha-helical regions and 12 beta-strands that together form three layers of sheet. The structure of the internal aldimine demonstrates that the PLP-cofactor is held in place by residues contributed from both subunits (Asp 164 and Gln 167 from Subunit I and Tyr 221 and Asn 235 from Subunit II). When the ketimine intermediate is present in the active site, the loop defined by Gln 225 to Ser 228 from Subunit II closes down upon the active site. The structure of DesV is similar to another sugar-modifying enzyme referred to as PseC. This enzyme is involved in the biosynthesis of pseudaminic acid, which is a sialic acid-like nonulosonate found in the flagellin of Helicobacter pylori. In the case of PseC, however, the amino group is transferred to the C-4 rather than the C-3 position. Details concerning the structural analysis of DesV and a comparison of its molecular architecture to that of PseC are presented.