Post-Translationally Pyroglutamate Formation on Proteins: Studies on protein production, crystal structures, catalysis mechanism, inhibition and pathophysiology of human glutaminyl cyclase
Date Issued
2006
Date
2006
Author(s)
Huang, Kai-Fa
DOI
en-US
Abstract
N-terminal pyroglutamate (pGlu) formation from its glutaminyl (or glutamyl) precursor is required in the maturation of numerous hormones and bioactive peptides. The aberrant formation of pGlu may be related to several pathological processes, such as osteoporosis, amyloidotic diseases, and rheumatoid arthritis. This N-terminal cyclization reaction, once thought to proceed spontaneously, was found to be catalyzed by the enzyme glutaminyl cyclase (QC). In this dissertation, the expression of functional human QC in Escherichia coli system to a relatively high level and to near homogeneity using a convenient purification procedure is present. We demonstrated, for the first time, that human QC contains one zinc ion per protein molecule. The enzyme was shown to be abundant in the brain tissues of normal adults and some patients with Alzheimer’s disease. This finding suggests that QC is responsible for the formation of N-terminal pGlu on several amyloid-related peptides, strengthening the possibility that human QC can be a drug target for treatment of several amyloidotic disorders. Furthermore, the first high-resolution crystal structures of human QC in free form and bound to a substrate and three imidazole-derived inhibitors are reported. The structure reveals an alpha/beta scaffold akin to that of two-zinc exopeptidases but with several insertions and deletions, particularly in the active-site region. The relatively closed active site of human QC displays alternate conformations primarily due to the different indole orientations of Trp207, resulting in two substrate (glutamine t-butyl ester)-binding modes. The single zinc ion in the active site is coordinated to three conserved residues and one water molecule, which is replaced by an imidazole nitrogen upon binding of the inhibitors. Together with structural and kinetic analyses of several active-site-mutant enzymes, a catalysis mechanism of the formation of protein N-terminal pGlu is proposed. These results provide a structural basis for the rational design of potent inhibitors of human QC.
Subjects
人類麩酸胺環化酵素
焦麩氨酸
後轉譯修飾作用
X─光繞射晶體學
阿茲海默症
鋅離子
human glutaminyl cyclase
pyroglutamate
post-translational modification
X-ray crystallography
zinc ion
SDGs
Type
other
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