A method of making a covalently coupled carrier polypeptide-target polypeptide conjugate, the method comprising: incorporating a first unnatural amino acid residue into a carrier polypeptide at a specific position during translation of the carrier polypeptide in a methylotrophic yeast cell, wherein the carrier polypeptide is a mammalian polypeptide and wherein incorporating the first unnatural amino acid into the carrier polypeptide at a specific position during translation comprises: (a) providing a methylotrophic yeast cell comprising (i) the first unnatural amino acid, (ii) an Escherichia coli orthogonal tRNA-synthetase (O-RS); (iii) an Escherichia coli orthogonal tRNA (O-tRNA), wherein the O-RS preferentially aminoacylates the O-tRNA with the first unnatural amino acid in the methylotrophic yeast cell; and (iv) a nucleic acid encoding the carrier polypeptide, wherein the nucleic acid comprises a selector codon that is recognized by the O-tRNA; and, (b) translating the nucleic acid, thereby incorporating the first unnatural amino acid into the carrier polypeptide during translation in the methylotrophic yeast cell; incorporating a second unnatural amino acid residue into a target polypeptide during synthesis or translation of the target polypeptide; and, reacting the first and second unnatural amino acid residues in vitro to produce the covalently coupled carrier polypeptide-target polypeptide conjugate. incorporating a first unnatural amino acid residue into a carrier polypeptide at a specific position during translation of the carrier polypeptide in a methylotrophic yeast cell, wherein the carrier polypeptide is a mammalian polypeptide and wherein incorporating the first unnatural amino acid into the carrier polypeptide at a specific position during translation comprises: (a) providing a methylotrophic yeast cell comprising (i) the first unnatural amino acid, (ii) an Escherichia coli orthogonal tRNA-synthetase (O-RS); (iii) an Escherichia coli orthogonal tRNA (O-tRNA), wherein the O-RS preferentially aminoacylates the O-tRNA with the first unnatural amino acid in the methylotrophic yeast cell; and (iv) a nucleic acid encoding the carrier polypeptide, wherein the nucleic acid comprises a selector codon that is recognized by the O-tRNA; and, (b) translating the nucleic acid, thereby incorporating the first unnatural amino acid into the carrier polypeptide during translation in the methylotrophic yeast cell; (a) providing a methylotrophic yeast cell comprising (i) the first unnatural amino acid, (ii) an Escherichia coli orthogonal tRNA-synthetase (O-RS); (iii) an Escherichia coli orthogonal tRNA (O-tRNA), wherein the O-RS preferentially aminoacylates the O-tRNA with the first unnatural amino acid in the methylotrophic yeast cell; and (iv) a nucleic acid encoding the carrier polypeptide, wherein the nucleic acid comprises a selector codon that is recognized by the O-tRNA; and, (i) the first unnatural amino acid, (ii) an Escherichia coli orthogonal tRNA-synthetase (O-RS); (iii) an Escherichia coli orthogonal tRNA (O-tRNA), wherein the O-RS preferentially aminoacylates the O-tRNA with the first unnatural amino acid in the methylotrophic yeast cell; and (iv) a nucleic acid encoding the carrier polypeptide, wherein the nucleic acid comprises a selector codon that is recognized by the O-tRNA; and, (b) translating the nucleic acid, thereby incorporating the first unnatural amino acid into the carrier polypeptide during translation in the methylotrophic yeast cell; incorporating a second unnatural amino acid residue into a target polypeptide during synthesis or translation of the target polypeptide; and, reacting the first and second unnatural amino acid residues in vitro to produce the covalently coupled carrier polypeptide-target polypeptide conjugate.