Aldose reductase from human psoas muscle. Affinity labeling of an active site lysine by pyridoxal 5'-phosphate and pyridoxal 5'-diphospho-5'-adenosine

The reaction of aldose reductase from human psoas muscle with either pyridoxal 5'-phosphate (PLP) or pyridoxal 5'-diphospho-5'-adenosine (PLP-AMP) results in a pseudo first-order 2-fold activation of the enzyme with the stoichiometric incorporation of 1 mol of either reagent per mol o...

Full description

Saved in:
Bibliographic Details
Published in:The Journal of biological chemistry 1989-02, Vol.264 (5), p.2912-2919
Main Authors: Morjana, N A, Lyons, C, Flynn, T G
Format: Article
Language:English
Subjects:
Adenosine Diphosphate - analogs & derivatives
Affinity Labels - metabolism
aldehyde reductase
Aldehyde Reductase - metabolism
Amino Acid Sequence
Binding Sites
Humans
Kinetics
Ligands
Male
Muscles - enzymology
Peptide Fragments - isolation & purification
Pyridoxal Phosphate - analogs & derivatives
Pyridoxal Phosphate - metabolism
skeletal muscle
Spectrophotometry
Sugar Alcohol Dehydrogenases - metabolism
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:The reaction of aldose reductase from human psoas muscle with either pyridoxal 5'-phosphate (PLP) or pyridoxal 5'-diphospho-5'-adenosine (PLP-AMP) results in a pseudo first-order 2-fold activation of the enzyme with the stoichiometric incorporation of 1 mol of either reagent per mol of enzyme. However, in addition to an increase in Vmax there was also an increase in Km for both substrate, DL-glyceraldehyde, and coenzyme, NADPH. This resulted in an overall decrease in catalytic efficiency (kcat/Km). Spectral analysis indicated that activation by both PLP and PLP-AMP was accompanied by Schiff's base formation and epsilon-pyridoxyllysine was identified in hydrolysates of the reduced enzyme PLP-complex. Digestion of either PLP-modified or PLP-AMP-modified aldose reductase with endoproteinase Lys-C followed by high performance liquid chromatography purification and amino acid sequencing of the pyridoxyllated peptide revealed that PLP and PLP-AMP had modified the same lysine residue. A 32-residue peptide containing the essential lysine was found to be highly homologous with a segment of the sequence of both human liver aldehyde reductase and rat lens aldose reductase. A tetrapeptide (Ile-Pro-Lys-Ser) containing the essential lysine was identical in all three enzymes. These results highlight the close structural similarity between members of the aldehyde reductase family.
ISSN:0021-9258
1083-351X