Native chemical ligation (NCL) has become the method of choice in synthesizing large or cyclic peptides/proteins. To overcome the limitation of NCL requiring N-terminal cysteine to mediate ligation, a strategy involving thiol-mediated ligation followed by desulfurization has been developed and advanced to realize peptide ligation at other amino acid sites, including Phe, Val, Leu, Thr, Lys, Pro and Gln. The syntheses of these mercapto-containing unnatural amino acids used as cysteine surrogates will be discussed in this review article.