Human trehalase: characterization, localization, and its increase in urine by renal proximal tubular damage.
By using polymerase chain reaction, cDNA encoding human renal trehalase has been isolated. The partial amino acid sequence deduced by the cDNA showed homologies in rabbit, Tenebrio molitor and silkworm trehalase. Northern blots showed renal trehalase mRNA to be about 2.0 kb. To examine the properties of renal and urinary human trehalase, the trehalase cDNA was inserted in the pMAL-cRI vector downstream from the malE gene, which encodes maltose-binding protein. Transfection of the recombinant pMAL-cRI in Escherichia coli provided high levels of expression of the maltose binding protein-trehalase fusion protein. A rabbit was immunized with purified fusion protein, and antihuman trehalase antibodies were obtained. Immunoblot analysis disclosed that renal and urinary trehalase exhibited a molecular mass of about 75 kDa. Analysis by indirect fluorescent microscopy demonstrated that the enzyme located in only proximal tubular cells. Urinary trehalase activity was low in the healthy infants and elevated in patients with asphyxia. Markedly high activity was observed in a patient with Lowe syndrome. The immunoreactive urinary trehalase with 75 kDa was increased dependent on the elevation of the activity. On the basis of these findings, we conclude that the increase of urinary trehalase reflects the extent of renal tubular damage, and we propose that urinary trehalase can be a specific marker of renal tubular damage.
Sasai-Takedatsu
M
M
Department of Pediatrics, Kansai Medical University, Osaka, Japan.
Taketani
S
S
Nagata
N
N
Furukawa
T
T
Tokunaga
R
R
Kojima
T
T
Kobayashi
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D015415:Biomarkers; D015152:Blotting, Northern; D012831:Bombyx; D003001:Cloning, Molecular; D018076:DNA, Complementary; D005198:Fanconi Syndrome / Q000201:enzymology* / Q000473:pathology / Q000652:urine; D006801:Humans; D007150:Immunohistochemistry; D007223:Infant; D007231:Infant, Newborn; D007687:Kidney Tubules, Proximal / Q000201:enzymology / Q000473:pathology; D008297:Male; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D011817:Rabbits; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid; D013712:Tenebrio; D014198:Trehalase / Q000096:biosynthesis / Q000737:chemistry* / Q000652:urine
D015415:Biomarkers; D018076:DNA, Complementary; D014198:Trehalase