This study investigates the microbial ecology of an Anaerobic Gas-Lift Reactor (AGR) to enhance biogas (methane) production from organic waste streams. Methods: Samples of vegetable waste, seed inoculum and food waste were collected from various locations in Hyderabad. Microbial isolation, characterization, and enzyme screening were conducted to identify key microbial species and their enzymatic capabilities. Growth dynamics under varying temperature and pH conditions were analyzed, and a microbial consortium was tested in a lab-scale biogas production system. Results: A diverse microbial community was identified, with extracellular enzyme screening revealing species with multifunctional enzymatic potential. Temperature and pH significantly influenced microbial growth and biogas production efficiency. Conclusion: The study provides valuable insights into optimizing microbial consortia for enhanced methane production, highlighting the critical roles of temperature and pH in regulating microbial activity. These findings contribute to developing efficient AGR systems for sustainable biogas generation from organic waste.