Journal
ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS
Volume 439, Issue 2, Pages 222-233Publisher
ELSEVIER SCIENCE INC
DOI: 10.1016/j.abb.2005.05.017
Keywords
monoterpene synthases; monoterpene cyclases; terpene cyclase structure-function; geranyl diphosphate cyclization; turpentine biosynthesis; directed mutagenesis; pinene; camphene; Abies grandis; Grand fir 3
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Funding
- NCI NIH HHS [R37 CA055254-14] Funding Source: Medline
- NIGMS NIH HHS [GM-31354] Funding Source: Medline
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Two monoterpene synthases, (-)-pinene synthase and (-)-camphene synthase, from grand fir (Abies grandis) produce different product mixtures despite having highly homologous amino acid sequences and, presumably, very similar three-dimensional structures. The major product of (-)-camphene synthase, (-)-camphene, and the major products of (-)-pinene synthase, (-)-alpha-pinene, and (-)-beta-pinene, arise through distinct mechanistic variations of the electrophilic reaction cascade that is common to terpenoid synthases. Structural modeling followed by directed mutagenesis in (-)-pinene synthase was used to replace selected amino acid residues with the corresponding residues from (-)-camphene synthase in an effort to identify the amino acids responsible for the catalytic differences. This approach produced an enzyme in which more than half of the product was channeled through an alternative pathway. It was also shown that several (-)-pinene synthase to (-)-camphene synthase amino acid substitutions were necessary before catalysis was significantly altered. The data support a model in which the collective action of many key amino acids, located both in and distant from the active site pocket, regulate the course of the electrophilic reaction cascade. (c) 2005 Elsevier Inc. All rights reserved.
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