Sidar Andika, Voshol Gerben P, El-Masoudi Ahmed, Vijgenboom Erik, Punt Peter J
Institute of Biology Leiden, Fungal Genetics and Biotechnology, Leiden University, 2333BE, Leiden, The Netherlands.
Department of Food and Agricultural Product Technology, Gadjah Mada University, Yogyakarta, 55281, Indonesia.
Fungal Biol Biotechnol. 2024 Sep 2;11(1):13. doi: 10.1186/s40694-024-00181-6.
Laccases are multi-copper oxidases that are usually composed of three Cu-oxidase domains. Domains one and three house the copper binding sites, and the second domain is involved in forming a substrate-binding cleft. However, Streptomyces species are found to have small laccases (SLAC) that lack one of the three Cu-oxidase domains. This type of SLAC with interesting lignocellulose bioconversion activities has not been reported in Aspergillus niger. In our research, we explored the expression and engineering of the SLAC from Streptomyces leeuwenhoekii C34 in A. niger. Genes encoding two versions of the SLAC were expressed. One encoding the SLAC in its native form and a second encoding the SLAC fused to two N-terminal CBM1 domains. The latter is a configuration also known for specific yeast laccases. Both SLAC variants were functionally expressed in A. niger as shown by in vitro activity assays and proteome analysis. Laccase activity was also analyzed toward bioconversion of lignocellulosic rice straw. From this analysis it was clear that the SLAC activity improved the efficiency of saccharification of lignocellulosic biomass by cellulase enzyme cocktails.
漆酶是多铜氧化酶，通常由三个铜氧化酶结构域组成。结构域一和结构域三含有铜结合位点，第二个结构域参与形成底物结合裂隙。然而，发现链霉菌属物种具有缺少三个铜氧化酶结构域之一的小型漆酶（SLAC）。这种具有有趣的木质纤维素生物转化活性的SLAC类型在黑曲霉中尚未见报道。在我们的研究中，我们探索了来自列文虎克链霉菌C34的SLAC在黑曲霉中的表达和工程改造。编码两种形式SLAC的基因被表达。一种编码天然形式的SLAC，另一种编码与两个N端CBM1结构域融合的SLAC。后者也是特定酵母漆酶所具有的一种构型。如体外活性测定和蛋白质组分析所示，两种SLAC变体均在黑曲霉中实现了功能性表达。还分析了漆酶对木质纤维素稻草生物转化的活性。从该分析中可以清楚地看出，SLAC活性提高了纤维素酶混合物对木质纤维素生物质的糖化效率。
