Isolation, identification and in silico analysis of alpha-amylase gene of Aspergillus niger strain CSA35 obtained from cassava undergoing spoilage

Abstract

In this investigation, a gene (CDF_Amyl) encoding extracellular α-amylase in Aspergillus niger strain CSA35 associated with cassava spoilage was amplified using specific primers and characterized in silico. The gene had a partial nucleotide sequence of 968 bp and encoded a protein of 222 aa residues with a molecular weight and isoelectric point of 25.13 kDa and 4.17, respectively. Its catalytic site was located in the active site domain. BLASTp analysis showed that the protein primary sequence of the α-amylase gene had 98% and 99% homologies with the α-amylase of A. niger and A. oryzae RIB40, respectively. The gene is more closely related to α-amylase genes from fungi than to bacterial, plant, or animal α-amylase genes. Restriction mapping of the gene showed it can be digested with restriction enzymes like NcoI, PstI, SmaI, and BcLI among others but not with EcoRI and EcoRV. Its protein product had a hydrophobicity score of − 0.43 but no transmembrane helix. The CDF_Amyl protein was subcellularly localized in the secretory pathway, an indication of its release into extracellular space after secretion. Also, the 3D structure of the CDF-Amyl protein was barrel-shaped with domains characteristic of α-amylases. The encoded α-amylase Vmax is 6.90 U/mg protein and Km is 6.70 mg/ml. It was concluded that the unique characteristics of the CDF_Amyl gene and its deduced protein could find applications in biotechnological, food and pharmaceutical industries where cloning and further modification of this gene would be required for product development and improvement.

Solid State Fermentation of Maize (<i>Zea mays</i>) Offal by <i>Rhizopus</i> <i>oligosporus</i> under Acidic and Basic Conditions

“…The enzyme activity was highest at fermentation pH 7 (509.04 ± 24.33 µg/g/min) when compared with the control. This may probably be due to the suitable environment created by the pH as well as activators of the enzyme present in the medium [19,20]. This report also agrees with the work of Aruna et al [21].…”

The Dual Prey-Inactivation Strategy of Spiders—In-Depth Venomic Analysis of Cupiennius salei

“…α-amylases are widely distributed in bacteria, archaea, fungi, and plants [33,34]. Moreover, many invertebrates and vertebrates express theses enzymes in their salivary glands and in the digestive liquid [35,36].…”

Microbial Alpha-Amylase Production: Progress, Challenges and Perspectives

“…), which cleaves internal alpha 1-4 glycosidic bands of starch and other polysaccharides to produce several products such as glucose and maltose. 7,8 It belongs to the family of GH13 (most of them), GH57, GH119, and GH126 9 and is one of the most widely used commercial enzymes. 10 Most of alpha-amylases are secreted extracellularly, however some intracellular alpha-amylase have been reported.…”