TY - JOUR
T1 - Characterization of His-tagged rat uroporphyrinogen III synthase wild-type and variant enzymes
AU - Li, Nan
AU - Ma, Dik Lung
AU - Liu, Xiaojun
AU - Wu, Long
AU - Chu, Xiusheng
AU - Wong, Kwok Yin
AU - Li, Ding
N1 - Funding Information:
Acknowledgments The work described in this paper was financially supported by the City University of Hong Kong.
PY - 2007/12
Y1 - 2007/12
N2 - The structurally related tetrapyrrolic pigments are a group of natural products that participate in many of the fundamental biosynthetic and catabolic processes of living organisms. Urogen III synthase catalyzes a key step in the formation of urogen III, a common intermediate for tetrapyrrolic natural products. In the present study, we cloned, purified, and characterized His-tagged rat urogen III synthase. The mechanism of enzymatic reaction was studied through site-directed mutagenesis of eight highly conserved residues with functional side chains around the active site followed with activity tests. Lys10, Asp17, Glu68, Tyr97, Asn121, Lys147, and His173 have not been studied previously, which were found to be unessential for enzymatic reaction. Tyr168 was identified as an important residue for enzymatic reaction catalyzed by rat urogen III synthase. Molecular modeling suggests the hydroxyl group of Tyr168 side chain is 3.5 Å away from the D ring, and is within hydrogen bond distance (1.9 Å) with acetate side chain of the D ring.
AB - The structurally related tetrapyrrolic pigments are a group of natural products that participate in many of the fundamental biosynthetic and catabolic processes of living organisms. Urogen III synthase catalyzes a key step in the formation of urogen III, a common intermediate for tetrapyrrolic natural products. In the present study, we cloned, purified, and characterized His-tagged rat urogen III synthase. The mechanism of enzymatic reaction was studied through site-directed mutagenesis of eight highly conserved residues with functional side chains around the active site followed with activity tests. Lys10, Asp17, Glu68, Tyr97, Asn121, Lys147, and His173 have not been studied previously, which were found to be unessential for enzymatic reaction. Tyr168 was identified as an important residue for enzymatic reaction catalyzed by rat urogen III synthase. Molecular modeling suggests the hydroxyl group of Tyr168 side chain is 3.5 Å away from the D ring, and is within hydrogen bond distance (1.9 Å) with acetate side chain of the D ring.
KW - Congenital erythropoetic porphyria
KW - spiro mechanism
KW - Tetrapyrrole
KW - Urogen III synthase
KW - Uroporphyrinogen III synthase
UR - http://www.scopus.com/inward/record.url?scp=35848970354&partnerID=8YFLogxK
U2 - 10.1007/s10930-007-9099-7
DO - 10.1007/s10930-007-9099-7
M3 - Journal article
C2 - 17763925
AN - SCOPUS:35848970354
SN - 1572-3887
VL - 26
SP - 569
EP - 576
JO - Protein Journal
JF - Protein Journal
IS - 8
ER -