Utility of Aspergillus niger citrate synthase promoter for heterologous expression

• Published in: Journal of biotechnology Vol. 155; no. 2; pp. 173 - 177
• Main Authors: Dave, Kashyap, Punekar, Narayan S.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 10.09.2011; Elsevier
• Subjects: Acetates; Aminobutyrates; Aspergillus niger; Aspergillus niger - enzymology; Aspergillus niger - genetics; Aspergillus niger - metabolism; Base Sequence; Biological and medical sciences; Biotechnology; carbon; citrate (si)-synthase; Citrate (si)-Synthase - genetics; Citrate (si)-Synthase - metabolism; Citrate synthase; DNA; Fundamental and applied biological sciences. Psychology; Fungal promoter; fungi; Gene constructs; genes; glufosinate; Glycerol; Green Fluorescent Proteins - metabolism; Medicin och hälsovetenskap; metabolism; Microscopy, Fluorescence; Molecular Sequence Data; mycelium; Mycelium - metabolism; Promoter Regions, Genetic - genetics; Protein expression; proteins; Sequence Analysis, DNA; Transformation, Genetic - genetics; tricarboxylic acid cycle; Protein expression; Fungal promoter; Gene constructs; Citrate synthase; Aspergillus niger; Enzyme; Citrate; Synthase; Gene expression; Protein; Fungi; Gene; Fungi Imperfecti
• ISSN: 0168-1656; 1873-4863; 1873-4863
• DOI: 10.1016/j.jbiotec.2011.06.012

Citrate synthase is a central player in the acidogenic metabolism of Aspergillus niger. The 5′ upstream sequence (0.9 kb DNA) of citrate synthase gene ( citA) from A. niger NCIM 565 was analyzed and its promoter function demonstrated through the heterologous expression of two proteins. The cloned citrate synthase promoter ( PcitA) sequence was able to express bar coding sequence thereby conferring phosphinothricin resistance. This sequence was further analyzed by systematic deletions to define an effective but compact functional promoter. The PcitA driven egfp expression showed that PcitA was active in all differentiation cell-stages of A. niger. EGFP expression was highest on non-repressible carbon sources like acetate and glycerol. Mycelial EGFP levels increased during acidogenic growth suggesting that PcitA is functional throughout this cultivation. A. niger PcitA is the first Krebs cycle gene promoter used to express heterologous proteins in filamentous fungi.