AbstractUnavoidable cracks cause a significant reduction in the strength and longevity of concrete. Water and several harmful ions seep through the cracks, initiate corrosion of the reinforcement, and affect the self-life of concrete. Self-repaired concrete will stand for a longer period and thus is gaining interest for constructions purposes. This work was an attempt to design a microbial protein (∼28 kDa) that incorporated self-healing cementitious material for future construction needs. The protein was isolated from an alkaliphilic hot spring bacterium (BKH4) of Bakreshwar, West Bengal, India. The prepared control and protein-amended cementitious mortar samples were subjected to simulate cracks and cured under water for several days. Images and microstructures of the control and protein-incorporated samples were analyzed, which established that there was a tiny fingers-like crystalline substance developed on the cracked surfaces. The developed substance was identified as a silicate phase (Gehlenite) by energy dispersive X-ray spectroscopic analysis. The microbial protein enhanced the mechanical strengths and durability of the protein-incorporated samples that were supported by the increments of ultrasonic pulse velocity, compressive strength, and sulfate resistance as well as reduction of water permeability and slow water movement (sorptivity test) of the experimental samples. This self-healing phenomenon is eco-efficient and developed due to the bio-silicification action of the microbial protein that was incorporated in mortar samples.