Molecular Evolution of the Substrate Specificity of Chloroplastic Aldolases/Rubisco Lysine Methyltransferases in Plants

Abstract

Rubisco and fructose-1,6-bisphosphate aldolases (FBAs) are involved in CO2 fixation in chloroplasts. Both enzymes are trimethylated at a specific lysine residue by the chloroplastic protein methyltransferase LSMT. Genes coding LSMT are present in all plant genomes but the methylation status of the substrates varies in a species-specific manner. For example, chloroplastic FBAs are naturally trimethylated in both Pisum sativum and Arabidopsis thaliana, whereas the Rubisco large subunit is trimethylated only in the former species. The in vivo methylation status of aldolases and Rubisco matches the catalytic properties of AtLSMT and PsLSMT, which are able to trimethylate FBAs or FBAs and Rubisco, respectively. Here, we created chimera and site-directed mutants of monofunctional AtLSMT and bifunctional PsLSMT to identify the molecular determinants responsible for substrate specificity. Our results indicate that the His-Ala/Pro-Trp triad located in the central part of LSMT enzymes is the key motif to confer the capacity to trimethylate Rubisco. Two of the critical residues are located on a surface loop outside the methyltransferase catalytic site. We observed a strict correlation between the presence of the triad motif and the in vivo methylation status of Rubisco. The distribution of the motif into a phylogenetic tree further suggests that the ancestral function of LSMT was FBA trimethylation. In a recent event during higher plant evolution, this function evolved in ancestors of Fabaceae, Cucurbitaceae, and Rosaceae to include Rubisco as an additional substrate to the archetypal enzyme. Our study provides insight into mechanisms by which SET-domain protein methyltransferases evolve new substrate specificity.

Identification of SET Domain-Containing Proteins in Gossypium raimondii and Their Response to High Temperature Stress

“…Ma et al . also found that LSMTs could trimethylate Rubisco in Fabaceae , Cucurbitaceae and Rosaceae , in addition to chloroplastic aldolases, which were only aldolases in most other plants10. However, possible biological functions of both GrS-ET and GrRBCMT proteins are still unclear in our current study.…”

“…KMT1 exhibits H3K9 substrate specificities activity, KMT2/KMT7 for H3K4, KMT3 for H3K36 and KMT6 for H3K27. RBCMT possesses H3K4 and H3K36 methyltransferase activity in animals, but non-histone target specific proteins in plant810. The function of S-ET is still unclear.…”

“…have recently proposed a new and rational nomenclature, in which plant SET domain-containing proteins were grouped into six distinct classes: KMT1 for H3K9, KMT2 for H3K4, KMT3 for H3K36, KMT6 for H3K27 and KMT7 for H3K4, while S-ETs contain an interrupted SET domain and are likely involved in the methylation of nonhistone proteins9. Besides the above major KMT classes, rubisco methyltransferase (RBCMT) family proteins are also identified as specific methyltransferases for nonhistone substrate in plants and consist of large subunit Rubisco methyltransferase (LSMT) and small subunit Rubisco methyltransferase (SSMT)810.…”

The thermostability and specificity of ancient proteins

“…One recent study found that an ancestral β-lactamase was both promiscuous and less efficient than its descendants [11]. Likewise, a study of RuBISCO found a promiscuous and inefficient ancestor, though this may be an artifact of poor reconstruction [35]. Other studies have determined the activities of ancient proteins, but not their specificity [6,7].…”

Protein lysine methylation contributes to modulating the response of sensitive and tolerant Arabidopsis species to cadmium stress

“…In leaf samples, where the immunodetection of the doublet of Lys‐methylated proteins is species and condition dependent (Figure a), we identified trimethylated Lys residues in EEF1A, RPL10A and six additional proteins (Table ). Methylation of chloroplastic fructose 1,6‐bisphosphate aldolases at a specific Lys residue (Lys395) was reported earlier (Alban et al, ; Ma et al, ; Mininno et al, ), while the other proteins were not previously known to be methylated.…”