Depending on the suboptimal factor, the target protein secretion can be over 1000-fold below the theoretical maximum. The bottlenecks may be alleviated by co-overexpression of "secretory helpers" (SHs). Here we proposed twelve SHs, functionally spanning the whole transcription-translation-translocation-folding-maturation-excretion pipeline. The genes were co-transformed with an easy-to-track reporter, and tested less than two temperatures. Our results indicated a clear distinction in the effects triggered by SHs involved in either synthesis or trafficking of the heterologous polypeptides. For superior operation of synthesis-related SHs, namely RPL3, SSA5 and SSA8, the secretory pathway's capacity must be released by applying decreased temperature (25 °C). The other SHs considered (e.g. SSO1, CWP11) did not give such spectacular results in the amounts of the target heterologous polypeptide, but allowed to maintain secretory capacity under unfavorable thermal conditions. This study provides generalizable guidelines for cloning/culturing strategies aiming at enhancement of heterologous protein secretion in Y. lipolytica.
Keywords: Protein trafficking; Secretion; Temperature; Transcription; Yeast.
© 2021 The Author(s).