The Del1 deposition domain can immobilize 3alpha-hydroxysteroid dehydrogenase in the extracellular matrix without interfering with enzymatic activity.
Developing methods that result in targeting of therapeutic molecules in gene therapies to target tissues has importance, as targeting can increase efficacy and decrease off target-side-effects. Work from my laboratory previously showed that the extracellular matrix protein Del1 is organized in the extracellular matrix (ECM) via the Del1 deposition domain (DDD). In this work, a fusion protein with DDD was made to assay the ability to immobilize an enzyme without disrupting enzymatic function. A prostatic cancer-derived cell line LNCap that grows in an androgen-dependent manner was used with 3alpha-hydroxysteroid dehydrogenase (3 alphaHD), which catalyzes dihydrotestosterone (DHT). Plasmids encoding a 3alphaHD:DDD fusion were generated and transfected into cultured cells. The effects of 3alphaHD immobilized in the ECM by the DDD were evaluated by monitoring growth of LNCap cells and DHT concentrations. It was demonstrated that the DDD could immobilize an enzyme in the ECM without interfering with function.
Hidai
Chiaki
C
Department of Biomedical Sciences, Nihon University School of Medicine, 30-1 Oyaguchikami-cho, Itabashi-ku, Tokyo, 173-8610, Japan. hidai@med.nihon-u.ac.jp
Kitano
Hisataka
H
Kokubun
Shinichiro
S
D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific) / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D015815:Cell Adhesion Molecules; D045744:Cell Line, Tumor; D004789:Enzyme Activation; D004800:Enzymes, Immobilized / Q000378:metabolism*; D005109:Extracellular Matrix / Q000378:metabolism*; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D008297:Male; D011471:Prostatic Neoplasms / Q000378:metabolism*
D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D015815:Cell Adhesion Molecules; C491706:Edil3 protein, mouse; D004800:Enzymes, Immobilized; D036341:Intercellular Signaling Peptides and Proteins; D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific)