Archaeal pyrrolysyl-tRNA synthetases (PylRSs) have been used to genetically encode over 200 distinct noncanonical amino acids (ncAAs) in proteins in and mammalian cells. This vastly expands the range of chemical functionality accessible within proteins produced in these organisms. Despite these clear successes, explorations of PylRS function in yeast remain limited. In this work, we demonstrate that the PylRS (MaPylRS) and its cognate tRNA support the incorporation of ncAAs into proteins produced in using stop codon suppression methodologies. Additionally, we prepared three MaPylRS mutants originally engineered in and determined that all three were active with one or more ncAAs, although with low efficiencies of ncAA incorporation in comparison to the parent MaPylRS. Alongside MaPylRS variants, we evaluated the activity of previously reported , , and chimeric and PylRSs. Using RJY100 and pairing these PylRSs with the tRNA, we did not observe any detectable stop codon suppression activity under the same conditions that produced moderately efficient ncAA incorporation with MaPylRS. The addition of MaPylRS/tRNA to the orthogonal translation machinery toolkit in potentially opens the door to hundreds of ncAAs that have not previously been genetically encodable using other aminoacyl-tRNA synthetase/tRNA pairs. Extending the scope of ncAA incorporation in yeast could powerfully advance chemical and biological research for applications ranging from basic biological discovery to enzyme engineering and therapeutic protein lead discovery.