Wax synthases (WS) catalyze the esterification of fatty alcohols and activated fatty acids to synthesize wax esters (WE). Enzymes with WS activities belong to different acyltransferase families with distinct properties. Some enzymes are not only monofunctional wax synthases but possess bi- or multifunctional activities. These enzymes are not only able to esterify fatty alcohols, but utilize diacylglycerols, diols, isoprenols or sterols as acyl-acceptors. This work has identified the first WS sequences from the eukaryotic protist Tetrahymena thermophila and from the preen glands of different bird species. The respective sequences were functionally expressed and characterized in yeast. These organisms were selected because they contain unusual wax ester species and might therefore contain WS enzymes with special substrate specificities. Via sequence homologies with human wax synthases we identified 4 proteins from Tetrahymena and 5 proteins from chicken which enabled the identification of two further sequences from goose and barn owl. Phylogenetic analysis of the sequences revealed that the Tetrahymena proteins as well as the avian WS4 and WS5 proteins build up own branches of acyl-CoA:diacylglycerol acyltransferase 2 (DGAT2) related proteins. GgWS1 and GgWS2 from chicken are more similar to mammalian DGAT2 family members. The proteins were analyzed via expression experiments in Saccharomyces cerevisiae mutants lacking TAG synthesis and via in vitro assay with membranes of transgenic yeast cells as enzyme source. These experiments demonstrated that all Tetrahymena thermophila (Tt) proteins were functionally expressed in S. cerevisiae and possessed both in vivo and in vitro acyltransferase activities. TtWS1 conferred mainly WS activity, whereas the other three TtWS proteins were multifunctional enzymes which also catalyzed the synthesis of triacylglycerol, isoprenyl esters and partly also diesters. Four out of five chicken sequences (Gallus gallus, Gg) were functionally expressed and resulted in the formation of storage lipids. GgWS2 and GgDGAT1 expressing yeast cultures produced wax esters, while GgWS1 and GgWS4 expressing cultures resulted in the additional accumulation of different amounts of triacylglycerol. Homologs of the chicken WS4 and WS5 from goose (Anser domesticus, Ad) and barn owl (Tyto alba, Ta) were cloned. The expression of both WS4 homologs resulted in the production of WE and TAG, while none of the WS5 homologs influenced lipid synthesis. In vitro experiments revealed that GgWS1 and GgWS2 were mainly monofunctional WS while the WS4 proteins belonged to the group of bifunctional WS/DGAT enzymes. AdWS4 and TaWS4 additionally showed high activities with isoprenols as acyl-acceptors in vitro. The comparison of lipid production in transgenic yeast cultures with lipids extracted from avian preen glands revealed different lipid compositions. Further in vitro experiments with membranes isolated from preen glands as enzyme source showed activites which were quite similar to the obtained results with yeast membranes. These experiments supported the idea that WE-species not only depend on the substrate specificities of the enzymes but also on the pool of accessible substrates in the tissues.