Generation of a Functionally Distinct Rhizopus oryzae Lipase through Protein Folding Memory

Rhizopus oryzae lipase (ROL) has a propeptide at its N-terminus that functions as an intramolecular chaperone and facilitates the folding of mature ROL (mROL). In this study, we successfully generated a functionally distinct imprinted mROL (mROL(imp)) through protein folding memory using a mutated propeptide. The mutated propeptide left its structural memory on mROL and produced mROL(imp) that exhibited different substrate specificities compared with mROL(WT) (prepared from the wild type propeptide), although the amino acid sequences of both mROLs were the same. mROL(imp) showed a preference for substrates with medium chain-length acyl groups and, noticeably, recognized a peptidase-specific substrate. In addition, ROL(imp) was more stable than mROL(WT). These results strongly suggest that proteins with identical amino acid sequences can fold into different conformations and that mutations in intramolecular chaperones can dynamically induce changes in enzymatic activity.