Laser welding experiments are performed on Ti-6Al-4V alloy sheets by adopting fiber laser. A special kind of workpiece fixture is designed and fabricated for providing shielding gas. After experiments, penetration depth, fusion zone width and heat-affected zone size at different locations within weld bead are measured and discussed in detail. Influence of line energies on the formation of non-uniform microstructure within weld bead is explored by conducting microstructural analysis. Various kinds of microstructural morphology of martensitic structure such as α′ martensite, blocky α, massive α and basket-weave microstructure are found in fusion zone. Experimentally, it is found that beam power is the key parameter for controlling penetration depth. Higher hardness is noticed within fusion zone due to the existence of large volume of α′ martensite. Tensile strength and hardness of welded specimens are increased with decreasing line energy. Small amount of micropores are also found in solidified weld bead. However, its sizes are in acceptable range as per BSEN:4678 standard. Most favorable welding condition is identified as a combination of beam power of 800 W and welding speed of 1000 mm/min which yields full penetration, narrower bead width, small heat-affected zone, minimal defects and acceptable mechanical properties.