8SWR

Structure of K. lactis PNP S42E variant bound to transition state analog DADMe-IMMUCILLIN G and sulfate

PDB DOI: https://doi.org/10.2210/pdb8SWR/pdb

Classification: TRANSFERASE
Organism(s): Kluyveromyces lactis NRRL Y-1140
Expression System: Escherichia coli BL21(DE3)
Mutation(s): Yes

Deposited: 2023-05-19 Released: 2023-10-18
Deposition Author(s): Fedorov, E., Ghosh, A.
Funding Organization(s): National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS), National Institutes of Health/National Cancer Institute (NIH/NCI)

Experimental Data Snapshot

Method: X-RAY DIFFRACTION
Resolution: 2.30 Å
R-Value Free: 0.236
R-Value Work: 0.201
R-Value Observed: 0.203

Starting Model: experimental
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wwPDB Validation 3D Report Full Report

Ligand Structure Quality Assessment

This is version 1.2 of the entry. See complete history.

Literature

Phosphate Binding in PNP Alters Transition-State Analogue Affinity and Subunit Cooperativity.

Minnow, Y.V.T., Schramm, V.L., Almo, S.C., Ghosh, A.

(2023) Biochemistry 62: 3116-3125

PubMed: 37812583 Search on PubMed
DOI: https://doi.org/10.1021/acs.biochem.3c00264
Primary Citation of Related Structures:
8SWP, 8SWQ, 8SWR, 8SWS, 8SWT, 8SWU

PubMed Abstract:
Purine nucleoside phosphorylases (PNPs) catalyze the phosphorolysis of 6-oxypurine nucleosides with an HPO ₄ ^2- dianion nucleophile. Nucleosides and phosphate occupy distinct pockets in the PNP active site. Evaluation of the HPO ₄ ^2- site by mutagenesis, cooperative binding studies, and thermodynamic and structural analysis demonstrate that alterations in the HPO ₄ ^2- binding site can render PNP inactive and significantly impact subunit cooperativity and binding to transition-state analogue inhibitors. Cooperative interactions between the cationic transition-state analogue and the anionic HPO ₄ ^2- nucleophile demonstrate the importance of reforming the transition-state ensemble for optimal inhibition with transition-state analogues. Altered phosphate binding in the catalytic site mutants helps to explain one of the known lethal PNP deficiency syndromes in humans.

Organizational Affiliation

Department of Biochemistry, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, New York 10461, United States.

Macromolecule Content

Total Structure Weight: 203.19 kDa
Atom Count: 13,130
Modelled Residue Count: 1,677
Deposited Residue Count: 1,848
Unique protein chains: 1

Macromolecules

Find similar proteins by:

(by identity cutoff) | 3D Structure

Entity ID: 1
Molecule	Chains	Sequence Length	Organism	Details	Image
Purine nucleoside phosphorylase	A, B, C, D, E A, B, C, D, E, F	308	Kluyveromyces lactis NRRL Y-1140	Mutation(s): 1 Gene Names: KLLA0_C16621g EC: 2.4.2.1
UniProt
Find proteins for Q6CSZ6 (Kluyveromyces lactis (strain ATCC 8585 / CBS 2359 / DSM 70799 / NBRC 1267 / NRRL Y-1140 / WM37)) Explore Q6CSZ6 Go to UniProtKB: Q6CSZ6
Entity Groups
Sequence Clusters	30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt Group	Q6CSZ6
Sequence Annotations Expand
Reference Sequence

Small Molecules

Ligands 3 Unique
ID	Chains	Name / Formula / InChI Key	2D Diagram	3D Interactions
IM5 (Subject of Investigation/LOI) Query on IM5 Download Ideal Coordinates CCD File SDF format, chain G [auth A] SDF format, chain I [auth B] SDF format, chain J [auth D] MOL2 format, chain G [auth A] MOL2 format, chain I [auth B] MOL2 format, chain J [auth D] mmCIF format, chain G [auth A] mmCIF format, chain I [auth B] mmCIF format, chain J [auth D]	G [auth A], I [auth B], J [auth D]	2-amino-7-{[(3R,4R)-3-hydroxy-4-(hydroxymethyl)pyrrolidin-1-yl]methyl}-3,5-dihydro-4H-pyrrolo[3,2-d]pyrimidin-4-one C₁₂ H₁₇ N₅ O₃ GSPTUGDLYPMLCQ-SFYZADRCSA-N		Interactions Focus chain G [auth A] Focus chain I [auth B] Focus chain J [auth D] Interactions & Density Focus chain G [auth A] Focus chain I [auth B] Focus chain J [auth D]
GUN Query on GUN Download Ideal Coordinates CCD File SDF format, chain L [auth E] MOL2 format, chain L [auth E] mmCIF format, chain L [auth E]	L [auth E]	GUANINE C₅ H₅ N₅ O UYTPUPDQBNUYGX-UHFFFAOYSA-N		Interactions Focus chain L [auth E] Interactions & Density Focus chain L [auth E]
SO4 (Subject of Investigation/LOI) Query on SO4 Download Ideal Coordinates CCD File SDF format, chain H [auth A] SDF format, chain K [auth D] MOL2 format, chain H [auth A] MOL2 format, chain K [auth D] mmCIF format, chain H [auth A] mmCIF format, chain K [auth D]	H [auth A], K [auth D]	SULFATE ION O₄ S QAOWNCQODCNURD-UHFFFAOYSA-L		Interactions Focus chain H [auth A] Focus chain K [auth D] Interactions & Density Focus chain H [auth A] Focus chain K [auth D]

Experimental Data & Validation

Experimental Data

Method: X-RAY DIFFRACTION
Resolution: 2.30 Å
R-Value Free: 0.236
R-Value Work: 0.201
R-Value Observed: 0.203
Space Group: P 1 2₁ 1

Unit Cell:

Length ( Å )	Angle ( ˚ )
a = 89.43	α = 90
b = 111.005	β = 108.3
c = 97.87	γ = 90

Software Package:

Software Name	Purpose
PHENIX	refinement
HKL-3000	data scaling
CBASS	data collection
MOSFLM	data reduction
PHASER	phasing

Structure Validation

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Ligand Structure Quality Assessment

Entry History & Funding Information

Deposition Data

Released Date: 2023-10-18

Deposition Author(s):

Fedorov, E.

Ghosh, A.

Funding Organization	Location	Grant Number
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)	United States	GM041916
National Institutes of Health/National Cancer Institute (NIH/NCI)	United States	CA013330