In the present work, cold rolling and cryo-rolling were performed on 99% commercially pure copper substrates. Both cold and cryo-rolling processes caused severe plastic deformation that led to an increase in dislocation density by 14× and 28× respectively, as compared to the pristine material. Increases in average tensile strengths, by 75% (488 MPa) and 150% (698 MPa), were observed in the two rolled materials as the result of the enhancement in dislocation density. In addition to strength, enhanced antibacterial property of cryo-rolled copper was observed in comparison to cold rolled and pristine copper. Initial adhesion and subsequent proliferation of bio-film forming Gram-positive bacteria Staphylococcus aureus was reduced by 66% and 100% respectively for cryo-rolled copper. Approximately 55% protein leakage, as well as ethidium bromide (EtBr) uptake, were observed confirming rupture of cell membrane of S. aureus. Inductively coupled plasma-mass spectroscopy reveals higher leaching of elemental copper in nutrient broth media from the cryo-rolled copper. Detailed investigations showed that increased dislocation led to leaching of copper ions that caused damage to the bacterial cell wall and consequently killing of bacterial cells. Cryo-rolling enhanced both strength, as well as antibacterial activity, due to the presence of dislocations. © 2018 by the authors.