Structural and functional studies of S-adenosyl-L-methionine binding proteins: a ligand-centric approach

Abstract

BackgroundThe post-genomic era poses several challenges. The biggest is the identification of biochemical function for protein sequences and structures resulting from genomic initiatives. Most sequences lack a characterized function and are annotated as hypothetical or uncharacterized. While homology-based methods are useful, and work well for sequences with sequence identities above 50%, they fail for sequences in the twilight zone (<30%) of sequence identity. For cases where sequence methods fail, structural approaches are often used, based on the premise that structure preserves function for longer evolutionary time-frames than sequence alone. It is now clear that no single method can be used successfully for functional inference. Given the growing need for functional assignments, we describe here a systematic new approach, designated ligand-centric, which is primarily based on analysis of ligand-bound/unbound structures in the PDB. Results of applying our approach to S-adenosyl-L-methionine (SAM) binding proteins are presented.ResultsOur analysis included 1,224 structures that belong to 172 unique families of the Protein Information Resource Superfamily system. Our ligand-centric approach was divided into four levels: residue, protein/domain, ligand, and family levels. The residue level included the identification of conserved binding site residues based on structure-guided sequence alignments of representative members of a family, and the identification of conserved structural motifs. The protein/domain level included structural classification of proteins, Pfam domains, domain architectures, and protein topologies. The ligand level included ligand conformations, ribose sugar puckering, and the identification of conserved ligand-atom interactions. The family level included phylogenetic analysis.ConclusionWe found that SAM bound to a total of 18 different fold types (I-XVIII). We identified 4 new fold types and 11 additional topological arrangements of strands within the well-studied Rossmann fold Methyltransferases (MTases). This extends the existing structural classification of SAM binding proteins. A striking correlation between fold type and the conformation of the bound SAM (classified as types) was found across the 18 fold types. Several site-specific rules were created for the assignment of functional residues to families and proteins that do not have a bound SAM or a solved structure.

Structural and Functional Studies of Pavine N-Methyltransferase from Thalictrum flavum Reveal Novel Insights into Substrate Recognition and Catalytic Mechanism

“…the Rossmann fold) is highly conserved (6,7). Structures of PavNMT in complex with SAM and SAH reveal a mode of cofactor binding that is most similar to Class Ia SAMdependent NMTs (7).…”

“…Three related NMTs with relatively distinct substrate specificities have been reported and include a coclaurine NMT (CNMT) involved in the formation of the branch-point intermediate (S)-reticuline (5); tetrahydroprotoberberine NMT (TNMT) (6), which participates in the formation of several BIAs, such as noscapine and sanguinarine; and pavine NMT (PavNMT) (7,8) (Fig. 1).…”

Systematic analysis of O -methyltransferase gene family and identification of potential members involved in the formation of O -methylated flavonoids in Citrus

“…Plant OMTs play important roles in the structural diversity of natural compounds. Enzymatic O-methylation of specific natural compounds changes their biological roles by altering their reactivity and solubility and their ability to interact with other chemical structures (Gana et al, 2013;Guidetti-Gonzalez et al, 2007).…”

“…and biotic stresses (Pichersky and Lewinsohn, 2011;Gana et al, 2013;Winkel-Shirley, 2001). Plant OMTs play important roles in the structural diversity of natural compounds.…”

Biosynthesis of wyosine derivatives in tRNAPhe of Archaea: role of a remarkable bifunctional tRNAPhe:m1G/imG2 methyltransferase

“…Spatial orientation of the bound AdoMet, usually in the (S,S) configuration (Cannon et al 2002), is of course crucial for the precise juxtaposition of the methyl group and the target C 7 -atom of imG-14 or N 1 -atom of guanine. Finally, a certain degree of flexibility of the target base within the anticodon loop of tRNA Phe (Sakaguchi et al 2012), the AdoMet (Gana et al 2013), or the mobility of certain amino acids of the active site of the enzyme (Christian et al 2006(Christian et al , 2013 may facilitate the electron donation along the methyl transfer process, probably by an S N 2 type of mechanism. If one accepts that certain Trm5-like enzymes may have a dual activity, it could be anticipated that after the emergence of the radical AdoMet enzyme Taw1, catalyzing the formation of tricyclic imG-14 from m 1 G-containing tRNA Phe , the evolving primordial cell might have benefited from the intrinsically relaxed specificity of its N 1 -methylating enzyme to also methylate the C 7 -atom of the imG-14 by a similar type of -atom of imG-14 is probably the same conserved E or D residue as mentioned above in the cases of other aTrm5, including MjTrm5b.…”