The putative amino-terminal signal peptide of the catalytic 19 kDa protein (KAZ) of Oplophorus luciferase was found to be a functional secretory peptide in mammalian cells. A 16 amino acid substituted mutant of KAZ (nanoKAZ) could be secreted from mammalian cells using the amino-terminal signal peptide of KAZ, but KAZ could not be secreted at all. Notably, nanoKAZ lacking the amino-terminal signal peptide could be secreted from mammalian cells, and the distribution of nanoKAZ on the cell membrane was confirmed by video-rate bioluminescence imaging. Thus, nanoKAZ lacking the amino-terminal signal peptide was expressed in the cytoplasm, translocated to the cell membrane, and released into the culture medium through an endoplasmic reticulum-Golgi-independent pathway. (C) 2014 Elsevier Inc. All rights reserved.