Lysosome-associated membrane protein-1 (LAMP-1) is a type I membrane glycoprotein consisting of a large luminal domain, a membrane-anchoring domain, and a short cytoplasmic tail (CT). The tyrosine-based motif (G³⁷⁸Y³⁷⁹QTI³⁸²) in its CT exclusively binds to adapter protein complex-3 (AP-3), which may facilitate the incorporation of LAMP-1 into transport vesicles to late endosomes and lysosomes. Of this sequence, Y³⁷⁹ is critical, and hydrophobic I³⁸² is optimal for the AP-3 binding and the efficient delivery to lysosomes. However, it is not clear how important G³⁷⁸ is in the AP-3 binding and lysosome transport. To clarify its importance, four mutants in which G³⁷⁸ was replaced with alanine, aspartic acid, glutamic acid, and asparagine (designated as G378A, G378D, G378E, and G378N, respectively) were prepared and their interaction strengths with AP-3 and lysosomal abundance were compared with those of wild-type (WT)-LAMP-1. A yeast two-hybrid system was applied to measure the interaction strengths of WT-, G378A-, G378D-, G378E-, and G378N-CTs with the medium subunit of AP-3 (μ3A). The G378A-, G378D-, and G378E-CTs as strongly interacted with μ3A as WT-CT, but the G378N-CT exhibited a very weak interaction with it. In the cell fractionation analyses, the lysosomal levels of G378A, G378D, and G378E were almost the same as that of WT whereas a lesser amount of G378N existed in the lysosomes. Taken together, it is considered that Y³⁷⁹ and I³⁸² are essential for AP-3-mediated vesicular transport of LAMP-1 while the Y³⁷⁹-preceding amino acid is not restricted to glycine but asparagine at this position is less suitable for that.