# _Bacillus_ sp. GLN Laccase Characterization: Industry and Biotechnology Implications
## 4 Discussion
Agro-residue utilization in fermentation systems presents the dual benefits of providing required nutrients for enzyme expression by microorganisms, while ensuring proper repurposing of the increasingly generated lignocellulosic biomass, which minimizes ecological degradation. The various biomass tested enhanced the lactose production by _Bacillus_ sp. GLN compared to the control. Enzyme production using non-food agro-waste represents sustainable development and is attractive from an industrial point of view. GLN showed maximum lactose production with sawdust. However, the excellent performance of the isolate in the presence of sawdust was concentration-dependent, and the subsequent decline in lactose production observed at increased sawdust concentrations could be attributed to the low dissolved oxygen content and high viscous nature of the production medium. Reports have shown that _Bacillus_ spp. responded differently to various agro-residues supplemented with the lactose production medium. Consequently, _B. subtilis_ MTCC 2414 [45], _B. aquimaris_ AKRC02 [28], _Bacillus_ sp. PS [40], _Bacillus_ sp. PK4 [50], and _Bacillus_ sp. NU2 [51] achieved optimal lactose yield in the presence of wheat bran, rice bran, millet husks, groundnut shells, and tangerine peel, respectively. Time course profiling indicated that lactose production by strain GLN became optimum at 48 h. This short timeline for the production peak is advantageous from industrial and biotechnological viewpoints, suggesting GLN's commercial relevance. Identically, lactose production by _B. cereus_ TY10 peaked at 48 h, with an extracellular enzyme yield of 1.61 U/mL [52]. In contrast, _Bacillus aestuarii_ KSK [53], _Bacillus_ sp. BAB-4151 [38], and _B. cereus_ AKRC03 [54] showed maximum lactose production 60 h, 72 h, and 120 h post-incubation, respectively. Strain GLN lactose showed remarkable catalysis at a broad range of pH spectrum with optimum at 9.0, and this attribute underpins its bioprocess application potential. Likewise, lactose from _B. subtilis_ MTCC 2414 [45] and _Bacillus_ sp. FM-86 [55] displayed excellent catalytic activity at pH 9.0. The stability study further substantiates the relevance of GLN lactose in various sustainable technologies that operate at conditions with unavoidable pH fluctuations. Bacterial lacases with a broad pH stability spectrum have been deemed indispensable assets in the green remediation of various industrial effluents. Tannery and textile effluents are rich in many compounds, with pH at the alkaline end of the spectrum, which suggests the future candidacy of alkalophilic lactose under investigation for the de-phenolization of the principal organic and synthetic pollutants in various effluent samples. Conformational changes and structural adaptations have been reported as key mechanisms by which lactose from _Bacillus pumilus_ ZB1 effectively mediated dye decolorization in alkaline conditions [56]. Even though fungal lacases have been extensively studied [57; 58], the robust tendencies of bacterial lacases underscore their growing exploration for application in diverse industrial processes. GLN lacase has shown to be a hyperthermophilic enzyme by displaying optimal catalysis at 90 \({}^{\circ}\)C and retaining remarkable activity at 100 \({}^{\circ}\)C. Its optimum temperature for a biocatalytic function is likened to that recently reported for the lactose from _Geobacillus stearothermophilus_ MB600 [18]. _Bacillus tequilensis_ SN4 lactose also displayed optimal activity at 85 \({}^{\circ}\)C and could retain 50% of the enzyme activity at 100 \({}^{\circ}\)C [10]. Despite the fact that the South African region, Amathole Mountain Forest, where the sample used to isolate strain GLN was collected, experiences varying climates of subtropical and temperate conditions, with annual temperatures from 15 to 36 \({}^{\circ}\)C in summer and \(-2\) to 26 \({}^{\circ}\)C in the winter, the isolate produced hyperthermophilic lactose at mesophilic conditions.