Recently, researchers have created novel fluorescent proteins by harnessing the somatic hypermutation ability of B cells. In this study, we examined if this approach could be used to evolve a non-fluorescent protein, namely the anti-apoptosis protein Bcl-x(L), using the Ramos B-cell line. After demonstrating that Ramos cells were capable of mutating a heterologous bcl-x(L) transgene, the cells were exposed to multiple rounds of the chemical apoptosis inducer staurosporine followed by rounds of recovery in fresh medium. The engineered B cells expressing Bcl-x(L) exhibited progressively lower increases in apoptosis activation as measured by caspase-3 activity after successive rounds of selective pressure with staurosporine treatment. Within the B-cell genome, a number of mutated bcl-x(L) transgene variants were identified after three rounds of evolution, including a mutation of Bcl-x(L) Asp29 to either Asn or His, in 8 out of 23 evaluated constructs that represented at least five distinct Ramos subpopulations. Subsequently, Chinese hamster ovary (CHO) cells engineered to overexpress the Bcl-x(L) Asp29Asn variant showed enhanced apoptosis resistance against an orthogonal apoptosis insult, Sindbis virus infection, when compared with cells expressing the wild-type Bcl-x(L) protein. These findings provide, to our knowledge, the first demonstration of evolution of a recombinant mammalian protein in a mammalian expression system.