Gene/Protein
Disease
Symptom
Drug
Enzyme
Compound
Pivot Concepts:
Gene/Protein
Disease
Symptom
Drug
Enzyme
Compound
Target Concepts:
Gene/Protein
Disease
Symptom
Drug
Enzyme
Compound
Query: EC:3.4.22.65 (
Der p 1
)
346
document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)
The mature cysteine protease from Dermatophgoides pteronyssinus,
Der p 1
, is a major house dust mite allergen. Its enzymatic activity has been shown to have pro-inflammatory effects that could also negatively influence efficacy of allergen-specific immunotherapy. The aim of this study was to express recombinant pro-
Der p 1
(rpro-
Der p 1
) in the yeast Pichia pastoris and to study its maturation. Expression was achieved at a concentration ranging from 45 mg.L-1 (
methanol
-induced expression) to 168 mg.L-1 (constitutive expression). No significant spontaneous maturation of the secreted proenzyme was observed. rpro-
Der p 1
with a sequence-based molecular mass of 34 kDa was hyperglycosylated by the yeast, migrating at 50-60 kDa on SDS/PAGE. Compared with its natural counterpart (nDer p 1), the recombinant proenzyme demonstrated decreased IgE reactivity, resulting in a 30-fold lower capacity to induce histamine release from human basophils. Decreased immunoreactivity was also shown by competitive RIA and sandwich ELISA with
Der p 1
-specific antibody reagents. CD spectra of rpro-
Der p 1
and nDer p 1 revealed significant structural differences. Deglycosylation of rpro-
Der p 1
with endoglycosidase H resulted in a decrease in apparent molecular mass from 50 kDa to 34 kDa, but did not affect nDer p 1. On removal of N-glycans from rpro-
Der p 1
, which harbours two putative N-glycosylation sites in both propeptide and mature sequence, the mature rDer p 1 appeared. This suggests that hyperglycosylation hampers spontaneous maturation. Maturation of the recombinant pro-enzyme was also achieved by addition of the active natural cysteine protease, nDer p 1. In conclusion, high-level expression of rpro-
Der p 1
in P. pastoris results in a stable hypoallergenic proenzyme with potential for use in allergen-specific immunotherapy.
...
PMID:Maturation of Pichia pastoris-derived recombinant pro-Der p 1 induced by deglycosylation and by the natural cysteine protease Der p 1 from house dust mite. 1185 27
