Isolation and Identification of Eukaryotic Initiation Factor 4A as a Molecular Target for the Marine Natural Product Pateamine A

Woon Kai Low, Yongjun Dang, Tilman Schneider-Poetsch, Zonggao Shi, Nam Song Choi, Robert M. Rzasa, Helene A. Shea, Shukun Li, Kaapjoo Park, Gil Ma, Daniel Romo, Jun Liu

Research output: Contribution to journalArticle

Abstract

Natural products continue to demonstrate their utility both as therapeutics and as molecular probes for the discovery and mechanistic deconvolution of various cellular processes. However, this utility is dampened by the inherent difficulties involved in isolating and characterizing new bioactive natural products, in obtaining sufficient quantities of purified compound for further biological studies, and in developing bioactive probes. Key to characterizing the biological activity of natural products is the identification of the molecular target(s) within the cell. The marine sponge-derived natural product Pateamine A (PatA) has been found to be an inhibitor of eukaryotic translation initiation. Herein, we describe the methods utilized for identification of the eukaryotic translation initiation factor 4A (eIF4A) as one of the primary protein targets of PatA. We begin by describing the synthesis of an active biotin conjugate of PatA (B-PatA), made possible by total synthesis, followed by its use for affinity purification of PatA binding proteins from cellular lysates. We have attempted to present the methodology as a general technique for the identification of protein targets for small molecules including natural products.

Original languageEnglish (US)
Pages (from-to)303-324
Number of pages22
JournalMethods in Enzymology
Volume431
DOIs
StatePublished - 2007

Fingerprint

Eukaryotic Initiation Factor-4A
Biological Products
Eukaryotic Initiation Factors
Molecular Probes
Porifera
Deconvolution
Biotin
Bioactivity
Purification
Carrier Proteins
Proteins
pateamine A
Molecules

ASJC Scopus subject areas

  • Biochemistry

Cite this

Isolation and Identification of Eukaryotic Initiation Factor 4A as a Molecular Target for the Marine Natural Product Pateamine A. / Low, Woon Kai; Dang, Yongjun; Schneider-Poetsch, Tilman; Shi, Zonggao; Choi, Nam Song; Rzasa, Robert M.; Shea, Helene A.; Li, Shukun; Park, Kaapjoo; Ma, Gil; Romo, Daniel; Liu, Jun.

In: Methods in Enzymology, Vol. 431, 2007, p. 303-324.

Research output: Contribution to journalArticle

Low, WK, Dang, Y, Schneider-Poetsch, T, Shi, Z, Choi, NS, Rzasa, RM, Shea, HA, Li, S, Park, K, Ma, G, Romo, D & Liu, J 2007, 'Isolation and Identification of Eukaryotic Initiation Factor 4A as a Molecular Target for the Marine Natural Product Pateamine A', Methods in Enzymology, vol. 431, pp. 303-324. https://doi.org/10.1016/S0076-6879(07)31014-8
Low, Woon Kai ; Dang, Yongjun ; Schneider-Poetsch, Tilman ; Shi, Zonggao ; Choi, Nam Song ; Rzasa, Robert M. ; Shea, Helene A. ; Li, Shukun ; Park, Kaapjoo ; Ma, Gil ; Romo, Daniel ; Liu, Jun. / Isolation and Identification of Eukaryotic Initiation Factor 4A as a Molecular Target for the Marine Natural Product Pateamine A. In: Methods in Enzymology. 2007 ; Vol. 431. pp. 303-324.
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