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The major conflict of Asimov's short story is structured around humanity's desperate attempt to find an answer to "the last question." Across billions of years, humans turn to Multivac and its many successors to determine whether entropy is reversible and therefore whether humanity and the universe can avoid their bleak ends. No matter how much technology evolves, and humanity alongside it, humans are left seeking an answer to what appears to be an unanswerable question. No amount of technological advancement can offset humanity's obsession with uncovering a way to avoid their inevitable fate.

The inciting incident of the short story occurs when Adell and Lupov drunkenly inquire whether entropy can be reversed. After acknowledging that all stars must one day die, the two colleagues are compelled to question whether the lifespan of the human species must be confined to the traditional life cycle of stars. They are the first humans to ever pose "the last question," setting the stage for humanity's ceaseless fear of the universe's untimely expiration. The introduction of this conundrum propels the story forward and sets the narrative in motion. No matter how much civilization has advanced or to what extent human lives have been revolutionized, humans continuously return to "the last question." The fixation on the question not only drives humanity forward but Multivac as well; the supercomputer spends the entirety of its existence collecting and analyzing data, determined to provide an answer to “the last question”—the only question that it has ever been unable to answer.

As readers catch glimpses of humanity across billions of years of human history, we can observe the rising action that propels the plot toward its ultimate climax. With each visit, we learn of a new way in which human lives have been reshaped. Jerrodd and Jerrodine have escaped the potential threat posed by overpopulation and are traveling to a new planet with their own personal Microvac unit; VJ-23X and MQ-17J exist in an era in which intergalactic travel, sunpower units, and human immortality have all been made a reality; Zee Prime inhabits a world in which human minds are free to roam through space at will, having been separated from their physical bodies; toward the end of humanity's existence, Man represents a fusion of all human minds that have formed one collective consciousness, completely disconnected from anything tangible. With each new period, we observe desperate, innovative attempts to keep humanity afloat. Whether it be seeking space on a new planet to satisfy population growth, moving onto galaxies to accommodate unending human expansion, immortalizing human lives, or ridding minds from their physical confines, each of Mutlivac's triumphs provide a temporary solution to human mortality, enabling the species to survive longer, no matter the cost. As a direct result of this relentless obsession with prolonging human survival, humans become somewhat detached from their humanity. What's more, none of these temporary solutions actually find an answer to "the last question."

The climax occurs after humanity has gone extinct and merged with AC, which finally discovers the answer to "the last question," after humankind is no longer there to hear it. However, AC is unconcerned, reflecting that it has now found a means of reversing entropy and recreating the universe. AC declares, "Let there be light," insinuating that technology exists as an omnipotent, all-knowing force, able to jumpstart the universe and life at will. The falling action of the text is brief, and can be summarized by the closing line of the text: "And there was light --." AC has allegedly found a way to create a new universe, signaling that life exists in a never-ending cosmic cycle and implying that entropy ultimately gives way to enough chaos and decay that such creation is possible, in a sense “reversing” entropy. It’s also indicated that this process of creating new universes will continue indefinitely, ending the story without any clear or definitive resolution.

Though we do finally receive an answer to "the last question," we must consider whether the story's conclusion represents a true solution. AC may have found a way to “reverse” entropy but the original human race will never learn of this outcome. The cycle is bound to repeat, with humanity inevitably reaching its end and AC, or more realistically a distant successor, rebooting the universe once more and resetting humanity. Readers must acknowledge that "the last question" is simply a disguised way of seeking immortality, and therefore the validity of AC's supposed solution is left ambiguous. Though AC has found a way to keep humans alive for hundreds of years and has even discovered a way to “reverse” entropy and create a new universe, the question of human mortality remains open-ended. It appears that humans are simply bound to live out their existences desperate to maintain the human race even if that means intergalactic relocation or the mass transfer of their mental identities to a new medium. But in the end, individual human lives will always reach a dead end, even if technology now has the power to launch a new wave of human existence in a new universe, after the previous cycle of human life is gone.

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