Gentamicin is an aminoglycoside antibiotic used in the treatment of gram-negative bacterial infections. However, its prolonged use is limited due to a high risk of nephrotoxicity. This study aimed to investigate the protective effects of a lipid transfer protein isolated from Morinda citrifolia L., Rubiaceae, seeds against gentamicin-induced nephrotoxicity in rats, and in a proximal tubular epithelial cell line from Rhesus monkey (LLC-MK2). Treatment with 6.25–25 μg/ml of the protein protected cells against cytotoxicity induced by 2 mM gentamicin. In the in vivo study, protein treatment (8 mg/kg, orally administered daily for 7 days 2 h after intraperitoneal injection of 100 mg/kg gentamicin) improved the levels of creatinine, urea, and uric acid. Additionally, it alleviated oxidative stress and inflammation induced by gentamicin, including a reduction in myeloperoxidase activity and cytokine modulation. Data were corroborated with histological analysis showing attenuated alterations by protein treatment. These findings suggest that this protein protects against gentamicin-induced nephrotoxicity. In addition, this natural product may be as an adjunctive tool to reduce kidney toxicity and improve the clinical usage profile of gentamicin.