Due to the low abundance of cysteine residues in proteins (<2%), significant efforts have been directed to link peptides with alternative amino acids. Desulfurization converts reactive cysteine thiol CH2SH to the CH3 group of alanine, which is significantly higher in abundance (>8%). Techniques include hydrogenation over Pd/Al2O3 and Raney nickel. Given the issues associated with purity, metal-free desulfurization alternative has been developed through a free-radical mechanism (Table , Entry 14; Figure ). However, native cysteines must be protected during desulfurization reactions. Acetamidomethyl cysteine (Cys(Acm)) is commonly employed in D-protein synthesis, and is commercially available in both L-and Denantiomers for Fmoc-SPPS (Figure ). Following desulfurization, the Acm group can be removed using mercury acetate, iodine, or more recently, PdCl2. Thiols may also be inserted into other canonical amino acids and can be removed by desulfurization (For review, see ). However, this approach has not been applied in D-protein synthesis because of challenges associated with synthesizing the enantiomeric building blocks (Table , Entry 15). Commercially available D-penicillamine (D-Pen) may be a viable option for ligation at valine (Figure ), but this residue is associated with slow ligation kinetics with all C-terminal thioester sites other than glycine. NCL can also proceed by employing a temporarily inserted thiol auxiliary which can be removed by acid cleavage. Such auxiliaries enable the generation of the lysine isopeptide bonds in the synthesis of branched ubiquitin chains (Table , Entry 25). Nevertheless, this approach has not yet been applied in the synthesis of the mirror-image D-counterparts (see section 3.3.2.3).