1G2O

CRYSTAL STRUCTURE OF PURINE NUCLEOSIDE PHOSPHORYLASE FROM MYCOBACTERIUM TUBERCULOSIS IN COMPLEX WITH A TRANSITION-STATE INHIBITOR

1G2O の概要

• エントリーDOI: 10.2210/pdb1g2o/pdb
• 分子名称: PURINE NUCLEOSIDE PHOSPHORYLASE, PHOSPHATE ION, 1,4-DIDEOXY-4-AZA-1-(S)-(9-DEAZAHYPOXANTHIN-9-YL)-D-RIBITOL, ... (4 entities in total)
• 機能のキーワード: trimer, transition-state complex, transferase
• 由来する生物種: Mycobacterium tuberculosis
• タンパク質・核酸の鎖数: 3
• 化学式量合計: 83882.04

構造登録者

Shi, W.,Basso, L.A.,Tyler, P.C.,Furneaux, R.H.,Blanchard, J.S.,Almo, S.C.,Schramm, V.L. (登録日: 2000-10-20, 公開日: 2001-08-01, 最終更新日: 2023-09-20)

主引用文献

Shi, W.,Basso, L.A.,Santos, D.S.,Tyler, P.C.,Furneaux, R.H.,Blanchard, J.S.,Almo, S.C.,Schramm, V.L.
Structures of purine nucleoside phosphorylase from Mycobacterium tuberculosis in complexes with immucillin-H and its pieces.
Biochemistry, 40:8204-8215, 2001

PubMed Abstract: A structural genomics comparison of purine nucleoside phosphorylases (PNPs) indicated that the enzyme encoded by Mycobacterium tuberculosis (TB-PNP) resembles the mammalian trimeric structure rather than the bacterial hexameric PNPs. The crystal structure of M. tuberculosis PNP in complex with the transition-state analogue immucillin-H (ImmH) and inorganic phosphate was solved at 1.75 A resolution and confirms the trimeric structure. Binding of the inhibitor occurs independently at the three catalytic sites, unlike mammalian PNPs which demonstrate negative cooperativity in ImmH binding. Reduced subunit interface contacts for TB-PNP, compared to the mammalian enzymes, correlate with the loss of the cooperative inhibitor binding. Mammalian and TB-PNPs both exhibit slow-onset inhibition and picomolar dissociation constants for ImmH. The structure supports a catalytic mechanism of reactant destabilization by neighboring group electrostatic interactions, transition-state stabilization, and leaving group activation. Despite an overall amino acid sequence identity of 33% between bovine and TB-PNPs and almost complete conservation in active site residues, one catalytic site difference suggests a strategy for the design of transition-state analogues with specificity for TB-PNP. The structure of TB-PNP was also solved to 2.0 A with 9-deazahypoxanthine (9dHX), iminoribitol (IR), and PO(4) to reconstruct the ImmH complex with its separate components. One subunit of the trimer has 9dHX, IR, and PO(4) bound, while the remaining two subunits contain only 9dHX. In the filled subunit, 9dHX retains the contacts found in the ImmH complex. However, the region of IR that corresponds to the oxocarbenium ion is translocated in the direction of the reaction coordinate, and the nucleophilic phosphate rotates away from the IR group. Loose packing of the pieces of ImmH in the catalytic site establishes that covalent connectivity in ImmH is required to achieve the tightly bound complex.

PubMed: 11444966
DOI: 10.1021/bi010585p

実験手法

X-RAY DIFFRACTION (1.75 Å)