Lactococcus lactis as expression host for the biosynthetic incorporation of tryptophan analogues in recombinant proteins

Incorporation of tryptophan (Trp) analogues into a protein may facilitate its structural analysis by spectroscopic techniques. Development of a biological system for the biosynthetic incorporation of such analogues into proteins is of considerable importance. The Gram-negative Escherichia coli is the only prokaryotic expression host regularly used for the incorporation of Trp analogues into recombinant proteins. Here we present the use of the versatile Gram-positive expression host Lactococcus lactis for the incorporation of Trp analogues. The availability of a tightly regulated expression system for this organism, the possibility to secrete modified proteins into the growth medium and the construction of the trp-synthetase deletion strain PA1002 of L. lactis rendered this organism potentially an efficient tool for the incorporation of Trp analogues in recombinant proteins. The Trp analogues 7-azatryptophan, 5-fluorotryptophan and 5-hydroxytryptophan were incorporated with efficiencies of >97%, >97% and 89%, respectively. Interestingly, 5-methyltryptophan could be incorporated with 92% efficiency. Successful biosynthetical incorporation of 5-methylTrp in recombinant proteins has not been reported before.