Application of affinity selection/mass spectrometry to determine the structural isomer of parnafungins responsible for binding polyadenosine polymerase.
To discover antifungal treatments that possess the desired characteristics of broad spectrum activity, a strong safety profile, and oral bioavailability, new discovery strategies must be implemented to identify structural classes of molecules capable of combating these microorganisms. One such technique that has been implemented is the Candida albicans Fitness Test, a whole cell screening platform capable of delineating the mechanism of action of compounds that demonstrate activity against the clinically relevant pathogenic fungus, C. albicans. Screening crude natural product extracts with this technology has resulted in the identification of a novel family of antifungal natural products, named the parnafungins, which inhibit the enzyme polyadenosine polymerase (PAP), a key component of the mRNA cleavage and polyadenylation complex. Owing to the rapid interconversion of the structural and stereoisomers of the parnafungins at neutral pH, the determination of the structural isomer with the highest affinity for PAP with standard biochemical assays has not been possible. Herein, we present an application of affinity-selection/mass spectrometry (AS-MS) to determine that the "straight" parnafungin structural isomer (parnafungin A) binds preferentially to PAP compared to the "bent" structural isomer (parnafungin B).
Adam
Gregory C
GC
Target Validation Chemistry, Natural Products Chemistry, and Infectious Diseases, Merck Research Laboratories, Merck and Company, P.O. Box 2000, Rahway, New Jersey 07065, USA. gregory_adam@merck.com
Parish
Craig A
CA
Wisniewski
Douglas
D
Meng
Juncai
J
Liu
Min
M
Calati
Kathleen
K
Stein
Benjamin D
BD
Athanasopoulos
John
J
Liberator
Paul
P
Roemer
Terry
T
Harris
Guy
G
Chapman
Kevin T
KT
D001688:Biological Products / Q000737:chemistry / Q000378:metabolism; D002853:Chromatography, Liquid; D005658:Fungi / Q000201:enzymology; D006801:Humans; D007536:Isomerism; D008024:Ligands; D013058:Mass Spectrometry; D023303:Oxazolidinones / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism*; D011062:Polynucleotide Adenylyltransferase / Q000378:metabolism*
D001688:Biological Products; D008024:Ligands; D023303:Oxazolidinones; D011062:Polynucleotide Adenylyltransferase