Protein S-palmitoylation refers to the covalent attachment of palmitic acid (C16:0) to the side chain of a cysteine residue on a protein through a thioester bond (Table 1). Palmitoylation is a reversible modification in organisms with specific enzymes to catalyze the addition or removal reaction [22,23], discovered in 1960s [24,25,26,27]. S-palmitoylation is dynamically regulated by palmitoyl acyltransferases (PATs, also known as ZDHHC-PATs) and acyl protein thioesterases (APTs), and the conserved zinc-finger Asp-His-His-Cys (ZDHHC) motif in the functional region of these enzymes is necessary for this process. PATs attach palmitoyl-CoA to proteins, and APTs perform depalmitoylation. For example, palmitic acid is added onto the SCRIB protein by the zinc finger DHHC-type palmitoyltransferase 7 (ZDHHC7) [28] and removed by lysophospholipase 2 (LYPLA2, also known as APT2) [29]. In some cases, however, proteins can be directly bound to palmitoyl-CoA and undergo PAT-independent auto-palmitoylation [30]. To date, a family of 24 mammalian PATs has been identified [7,31]. The “SwissPalm” database shows that > 10% of the human proteome is susceptible to S-palmitoylation [32,33], in which >600 substrates have already been fully or partially characterized [34,35,36]. Proteins that can be S-palmitoylated include NRas proto-oncogene, GTPase (NRAS) [37], β₂ adrenergic receptors (β₂ARs) [38], Fas cell surface death receptor (FAS) [39], BCL2-associated X, apoptosis regulator (BAX) [40], inositol 1,4,5-triphosphate receptor type I (ITPR1) [41], junctional adhesion molecule 3 (JAM3) [42], and SRC proto-oncogene, non-receptor tyrosine kinase (SRC) [43].

N-palmitoylation is classified into N-terminal palmitoylation and N^ε-palmitoylation according to the position of the modification in the protein (Table 1). In N-terminal palmitoylation, palmitic acid is linked to the amino group of the cysteine residue at the N-terminus of substrate proteins, whereas in N^ε-palmitoylation, palmitic acid is covalently attached to the ε-amino group of the lysine residue at the N-terminus via an amide bond. The biological significance of N-terminal palmitoylation has been reviewed before [31,44,74]. A unique dual palmitoylation in the N-terminal region of the human LIM domain kinase 1 (LIMK1) controls the targeting of this protein to the spine and contributes to the activation of the protein by membrane-localized p21-activated kinase (PAK) [74]. N-terminal palmitoylation has also been detected in Sonic Hedgehog (SHh) proteins [75] and shown to be catalyzed by Hedgehog acyltransferase (HHAT) [76]. Additionally, Sirtuin (SIRT) has been reported to be modified by N^ε-palmitoylation [45,46,77,78].

Palmitic acid can be irreversibly linked to the side chain of serine residues in proteins via ester bonds in organisms without specific enzymes removing the attached lipid chain (Table 1). Currently, only a few proteins are known to be O-palmitoylated. One of them is histone H4, which can be O-palmitoylated at Ser-45 by an enzyme called lysophosphatidylcholine acyltransfer ase 1 (LPCAT1) [47]. Interestingly, O-palmitoylation at the threonine residue in the C-terminal of the spider venom neurotoxin PLTX-II has been reported, and it is thought to regulate the toxin activity in blocking presynaptic voltage-gated Ca²⁺ channels [48].

Similar to N-palmitoylation, N-myristoylation is also categorized into two major classes—N-terminal myristoylation and N^ε-myristoylation (Table 1). N-terminal myristoylation is the attachment of myristic acid (C14:0) to a protein N-terminus with a glycine residue through an amide linkage. This reaction is catalyzed by N-myristoyltransferases (NMTs), which recognize the GXXXS/T signature sequence (where X is any amino acid) in substrate proteins [49]. Human proteomic studies have suggested that > 100 proteins are N-myristoylated [79], such as A-kinase anchoring protein 12 (AKAP12) [77], SRC [78], protein kinase AMP-activated non-catalytic subunit beta 1 (PRKAB1) [80], FMR1 autosomal homolog 2 (FXR2) [81], and hexokinase 1 variant in mammalian spermatozoa (HK1S) [82], underscoring the vital roles of N-myristoylation [83]. N^ε-myristoylation refers to the attachment of myristic acid to the ε-amino group of a lysine residue in substrate proteins. The enzyme(s) that catalyze this PTM is (are) not known; however, N^ε-myristoylated proteins can efficiently be deacylated by SIRTs [50,51,52].

In addition to palmitoylation and myristoylation, proteins can be lipidated with other types of long-, medium-, or short-chain saturated fatty acids (Table 1). Studies have shown that influenza virus hemagglutinin can be S-acylated by stearate (C18:0) [53,54], Ghrelin can be O-octanoylated (C8:0) [55,56]. In 2009, the Zhao group released a bioinformatic tool named PTMap [84], which helps identify various acylations, including Lys propionylation, butyrylation, hydroxyl-fatty acid modification, lactylation, bicarboxylic acid modification, and benzoylation. Numerous studies have discovered >500 histone modification sites for above listed modifications, greatly expanding the understanding of PTM and histone modifications [85].

Physeterylation (C14:1n9) is detected in the retina, heart, and liver [86] and on SRC family kinases [87]. Myristoleoyted (C14:1n5) proteins have also been found [88,89]. WNT proteins are O-palmitoleoylated with palmitoleic acid (C16:1n7) on their conserved serine residue by the O-acyltransferase Porcupine [57,58,90], and the palmitoleic acid of an O-palmitoleoylated WNT protein is removed by Notum [59]. Oleic acid (C18:1n9) modification has been reported on the lysine residue of the lens integral membrane protein aquaporin-0 and plays an important role in targeting the substrate protein to membrane domains in the bovine and human lens [60] (Table 1).