Aktivitas Enzim β-1,3-Glukanase dari Streptomyces spp. Rizosfer dan Kloning Gen Penyandinya

Abstract

Enzim β-1,3-glukanase merupakan salah satu enzim yang mampu menghidrolisis glukan penyusun dinding sel cendawan fitopatogen. Aktinomiset rizosfer jagung dari kelompok Streptomyces spp. diduga mampu memproduksi enzim yang memiliki aktivitas glukanolitik dan berpotensi untuk dikembangkan sebagai agen biokontrol cendawan fitopatogen. Penelitian ini bertujuan menganalisis aktivitas enzim glukanase secara kuantitatif dua isolat Streptomyces spp. dengan indeks glukanolitik tertinggi pada penelitian sebelumnya dan mengidentifikasi gen penyandinya. Metode penelitian yang dilakukan meliputi uji aktivitas glukanase secara kuantitatif, amplifikasi gen endo-β-1,3-glukanase (bglS), kloning, konfirmasi hasil kloning, sekuensing, analisis fenetik, dan prediksi model protein. Aktivitas spesifik enzim isolat S. collinus ARJ38 lebih tinggi dari S. tritolerans ARJ32, yakni sebesar 41,599 U/mg pada waktu inkubasi hari ke-8. Keberadaan gen penyandi bglS pada kedua isolat berhasil diamplifikasi dan dikloning. Hasil sekuensing gen bglS parsial dari kedua isolat potensial teridentifikasi sebagai endo-β-1,3-glukanase dari kelompok glycoside hydrolase (GH) family 16. Model struktur tiga dimensi yang dikonstruksi dari sekuens asam amino bglS parsial memiliki parameter kualitas serta kemiripan yang tinggi dengan model protein eksperimental endo-β-1,3-glukanase parsial dari Nocardiopsis sp. F96.

The β-1,3-glucanase enzyme is one of the enzymes capable of hydrolyzing the glucan components of phytopathogenic fungal cell walls. Streptomyces spp. isolated from the maize rhizosphere are suspected to produce enzymes with glucanolytic activity and have the potential to be developed as biological controlling agents against phytopathogenic fungi. This study aims to quantitatively analyze the enzymatic activity of glucanase in two Streptomyces spp. isolates with the highest glucanolytic index from previous research, and to identify their encoding genes. The research methods include quantitative analyze of the glucanase activity, amplification of the endo-β-1,3-glucanase (bglS) gene, cloning, confirmation of cloning results, sequencing, phenetic tree analysis, and protein model prediction. The specific activity enzyme of S. collinus ARJ38 was higher than that of S. tritolerans ARJ32, with a value of 41.599 U/mg on the 8th day of incubation. The presence of the bglS gene in both isolates was successfully amplified and cloned. The sequencing results of the bglS gene from both potential isolates were identified as endo-β-1,3-glucanase from the glycoside hydrolase family 16. The three-dimensional structure model constructed from the partial bglS amino acid sequence exhibited high-quality parameters and high similarity with the experimental protein model of partial endo-β-1,3-glucanase from Nocardiopsis sp. F96.