1. ¹Edward A. Doisy Department of Biochemistry and Molecular Biology, Saint Louis University School of Medicine, 1402 South Grand Boulevard, St. Louis, MO 63104, USA
2. ³Department of Pathology, Saint Louis University School of Medicine, 1402 South Grand Boulevard, St. Louis, MO 63104, USA
3. ²Department of Pediatrics, Gifu University School of Medicine, 40 Tsukasa-machi, Gifu 500-5715, Japan
4. ⁴Department of Internal Medicine, Program in Gene Therapy, University of Iowa College of Medicine, Iowa City, IA 52242, USA
5. ⁵Department of Human Welfare, Chubu Gakuin University, Kurachi, Seki, Gifu, Japan

Correspondence: William S. Sly, Fax: +1 (314) 977 1183. E-mail: slyws@slu.edu

Received 5 November 2004; Accepted 18 February 2005.

Abstract

We used recombinant forms of human -glucuronidase (GUS) purified from secretions from stably transfected CHO cells to compare the native enzyme to a GUS–Tat C-terminal fusion protein containing the 11-amino-acid HIV Tat protein transduction domain for: (1) susceptibility to endocytosis by cultured cells, (2) rate of clearance following intravenous infusion, and (3) tissue distribution and effectiveness in clearing lysosomal storage following infusion in the MPS VII mouse. We found: (1) Native GUS was more efficiently taken up by cultured human fibroblasts and its endocytosis was exclusively mediated by the M6P receptor. The GUS–Tat fusion protein showed only 30–50% as much M6P-receptor-mediated uptake, but also was taken up by adsorptive endocytosis through binding of the positively charged Tat peptide to cell surface proteoglycans. (2) GUS–Tat was less rapidly cleared from the circulation in the rat (t_1/2 = 13 min vs 7 min). (3) Delivery to most tissues of the MPS VII mouse was similar, but GUS–Tat was more efficiently delivered to kidney. Histology showed that GUS–Tat more efficiently reduced storage in renal tubules, retina, and bone. These studies demonstrate that Tat modification can extend the range of tissues corrected by infused enzyme.