Formylation

Formylation refers to any chemical processes in which a compound is functionalized with a formyl group ( CH=O). In organic chemistry, the term is most commonly used with regards to aromatic compounds (for example the conversion of benzene to benzaldehyde in the Gattermann–Koch reaction). In biochemistry the reaction is catalysed by enzymes such as formyltransferases. Formylation generally involves the use of formylation agents, reagents that give rise to the CHO group. Among the many formylation reagents, particularly important are formic acid and carbon monoxide. A formylation reaction in organic chemistry refers to organic reactions in which an organic compound is functionalized with a formyl group ( CH=O). The reaction is a route to aldehydes (C CH=O), formamides (N CH=O), and formate esters (O CH=O).

Formylation reactions are a form of electrophilic aromatic substitution and therefore work best with electron rich starting materials. Phenols are a common substrate, as they readily deprotonate to excellent phenoxide nucleophiles. Other electron rich substrates, such as mesitylene, pyrrole, or fused aromatic rings can also be expected to react. Benzene will react under aggressive conditions but deactivated rings such as pyridine are difficult to formylate effectively. Many formylation reactions will select only the ortho product (e. g. salicylaldehyde), attributed to attracton between the phenoxide and the formylating reagent. Ionic interactions have been invoked between cationic nitrogen centres in the Vilsmeier–Haack reaction and Duff reaction; coordination to high oxidation metals has been invoked in the Casiraghi and Rieche formylations (cf. Kolbe–Schmitt reaction). The direct reaction between phenol and paraformaldehyde is possible via the Casiraghi formylation, but other methods apply masked forms of formaldehyde, in part to limit the formation of phenol formaldehyde resins. Aldehydes are strongly deactivating and as such phenols typically only react once. However certain reactions, such as the Duff reaction, can give double addition. Formylation can be applied to other aromatic rings. As it generally begins with nucleophilic attack by the aromatic group, the electron density of the ring is an important factor. Some aromatic compounds, such as pyrrole, are known to formylate regioselectively. Formylation of benzene rings can be achieved via the Gattermann reaction and Gattermann Koch reaction. These involve strong acid catalysis and proceed in a manner similar to the Friedel–Crafts reaction.

Hydroformylation of alkenes is the most important method for obtaining aliphatic formyls (i. e., aldehydes). The reaction is largely restricted to industrial settings. Several specialty methods exist for laboratory scale synthesis, including the Sommelet reaction, Bouveault aldehyde synthesis or Bodroux–Chichibabin aldehyde synthesis.

In biochemistry, the addition of a formyl functional group is termed "formylation". A formyl functional group consists of a carbonyl bonded to hydrogen. When attached to an R group, a formyl group is called an aldehyde. Formylation has been identified in several critical biological processes. Methionine was first discovered to be formylated in E. coli by Marcker and Sanger in 1964 and was later identified to be involved in the initiation of protein synthesis in bacteria and organelles. The formation of N formylmethionine is catalyzed by the enzyme methionyl tRNA transformylase. Additionally, two formylation reactions occur in the de novo biosynthesis of purines. These reactions are catalyzed by the enzymes glycinamide ribonucleotide (GAR) transformylase and 5 aminoimidazole 4 carboxyamide ribotide (AICAR) transformylase. More recently, formylation has been discovered to be a histone modification, which may modulate gene expression.

Formylation of methanofuran initiates the methanogenesis cycle. The formyl group is derived from carbon dioxide and is converted to methane.

In bacteria and organelles, the initiation of protein synthesis is signaled by the formation of formyl methionyl tRNA (tRNAfMet). This reaction is dependent on 10 formyltetrahydrofolate, and the enzyme methionyl tRNA formyltransferase.

The formylation reaction is proposed to occur through a direct transfer reaction in which the amine group of GAR nucleophilically attacks N10 formyl THF creating a tetrahedral intermediate.

PurT GAR transformylase requires formate as the formyl donor and ATP for catalysis. It has been estimated that PurT GAR transformylase carries out 14 50% of GAR formylations in E. coli. The enzyme is a member of the ATP grasp superfamily of proteins.

A sequential mechanism has been proposed for PurT GAR transformylase in which a short lived formyl phosphate intermediate is proposed to first form. This formyl phosphate intermediate then undergoes nucleophilic attack by the GAR amine for transfer of the formyl group. A formyl phosphate intermediate has been detected in mutagenesis experiments, in which the mutant PurT GAR transforymylase had a weak affinity for formate. An enzyme phosphate intermediate preceding the formylphosphate intermediate has also been proposed to form based on positional isotope exchange studies. Formylation is detected most frequently on 19 different modification sites on Histone H1. The genetic expression of the cell is highly disrupted by formylation, which may cause diseases such as cancer. The development of these modifications may be due to oxidative stress. The formylphosphate produced can then be used to formylate lysine. Oxidative stress is believed to play a role in the availability of lysine residues in the surface of proteins and the possibility of being formylated.

Inhibition of enzymes involved in purine biosynthesis has been exploited as a potential drug target for chemotherapy. Cancer cells require high concentrations of purines to facilitate division Several folate based inhibitors have been developed to inhibit formylation reactions by GAR transformylase and AICAR transformylase. The first GAR transformylase inhibitor Lometrexol [(6R)5,10 dideazatetrahydrofolate] was developed in the 1980s through a collaboration between Eli Lilly and academic laboratories. Although similar in structure to N10 formyl THF, lometrexol is incapable of carrying out one carbon transfer reactions. Exome sequencing, has been used to identify a mutation in the gene coding for mitochondrial methionyl tRNA formyltransferase (MTFMT) in patients with Leigh syndrome. The c.626C>T mutation identified in MTFMT yielding symptoms of Leigh Syndrome is believed to alter exon splicing leading to a frameshift mutation and a premature stop codon. Individuals with the MTFMT c.626C>T mutation were found to have reduced fMet tRNAMet levels and changes in the formylation level of mitochondrically translated COX1. This link provides evidence for the necessity of formylated methionine in initiation of expression for certain mitochondrial genes.