2HCT

Acidic residues at the active sites of CD38 and ADP-ribosyl cyclase determine NAAPD synthesis and hydrolysis activities

2HCT の概要

• エントリーDOI: 10.2210/pdb2hct/pdb
• 分子名称: ADP-ribosyl cyclase 1, BETA-NICOTINAMIDE RIBOSE MONOPHOSPHATE (3 entities in total)
• 機能のキーワード: beta sheets, alpha bundle, hydrolase
• 由来する生物種: Homo sapiens (human)
• 細胞内の位置: Membrane; Single-pass type II membrane protein: P28907
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 61287.11

構造登録者

Liu, Q.,Kriksunov, I.A.,Hao, Q.,Graeff, R.,Lee, H.C. (登録日: 2006-06-18, 公開日: 2006-08-22, 最終更新日: 2024-11-13)

主引用文献

Graeff, R.,Liu, Q.,Kriksunov, I.A.,Hao, Q.,Lee, H.C.
Acidic residues at the active sites of CD38 and ADP-ribosyl cyclase determine nicotinic acid adenine dinucleotide phosphate (NAADP) synthesis and hydrolysis activities.
J.Biol.Chem., 281:28951-28957, 2006

PubMed Abstract: Nicotinic acid adenine dinucleotide phosphate (NAADP) is a novel metabolite of NADP that has now been established as a Ca(2+) messenger in many cellular systems. Its synthesis is catalyzed by multifunctional enzymes, CD38 and ADP-ribosyl cyclase (cyclase). The degradation pathway for NAADP is unknown and no enzyme that can specifically hydrolyze it has yet been identified. Here we show that CD38 can, in fact, hydrolyze NAADP to ADP-ribose 2'-phosphate. This activity was low at neutrality but greatly increased at acidic pH. This novel pH dependence suggests that the hydrolysis is determined by acidic residues at the active site. X-ray crystallography of the complex of CD38 with one of its substrates, NMN, showed that the nicotinamide moiety was in close contact with Glu(146) at 3.27 A and Asp(155) at 2.52 A. Changing Glu(146) to uncharged Gly and Ala, and Asp(155) to Gln and Asn, by site-directed mutagenesis indeed eliminated the strong pH dependence. Changing Asp(155) to Glu, in contrast, preserved the dependence. The specificity of the two acidic residues was further demonstrated by changing the adjacent Asp(147) to Val, which had minimal effect on the pH dependence. Crystallography confirmed that Asp(147) was situated and directed away from the bound substrate. Synthesis of NAADP catalyzed by CD38 is known to have strong preference for acidic pH, suggesting that Glu(146) and Asp(155) are also critical determinants. This was shown to be case by mutagensis. Likewise, using similar approaches, Glu(98) of the cyclase, which is equivalent to Glu(146) in CD38, was found to be responsible for controlling the pH dependence of NAADP synthesis by the cyclase. Based on these findings, a catalytic model is proposed.

PubMed: 16861223
DOI: 10.1074/jbc.M604370200

実験手法

X-RAY DIFFRACTION (1.95 Å)