In the year 2199, Earth is ravaged by attacks from alien beings of the the planet, Gamilus, which have devastated the surface of the Earth and forced what remains of mankind to take shelter underground. To save the Earth, the last of the Earth's space forces must distract the Gamilan forces while a non-Gamilan ship, the Amaterasu, lands on Mars to deliver the Wave Motion Activation Core to the Earthlings. While Captain Mamoru Kodai contains the enemy attack to allow fellow captain Juzo Okita to escape with his forces, Mamoru's brother Susumu and his friend Daisuke Shima investigate the Amaterasu that crashed on Mars. Inside the ship there is the corpse of an alien female, the messenger sent to Earth to make the important delivery to the Earthlings. Susumu and Daisuke return to Earth on the Kirishima, the only ship to return from the battle at Pluto. On the Kirishima, Susumu learns of his brother's death. When the Kirishima reaches Earth, Susumu Kodai looks for Captain Okita to know whether his brother was a decoy or not and meets Yuki Mori, who directs him to the hospital wing. Kodai confronts Okita, who says that it was he who led Mamoru to his death.

Susumu brings the Wave Motion Activation Core to Earth which allows mankind to complete its first interstellar spaceship, the "BBY-01 Yamato". With Okita as the captain, Shima as the navigator, and Susumu as its battle chief, the Yamato is assigned to travel to planet Iscandar, in the Great Magellanic Cloud, with orders to retrieve a device able to restore Earth's environment. Before the Yamato can start its journey, it must counter a Gamilan interplanetary ballistic missle aimed at them.

On Pluto, Gantz suggests sending out a fleet after the Yamato. Schultz turns down the suggestion, preferring to see what the Yamato does before attempting anything. After making a space warp, the Yamato 's main engine isn't transferring energy correctly, so Captain Okita orders the Yamato to land on a floating continent orbiting Jupiter, to make repairs to the engine. On the floating continent, Captain Okita orders the crew of the Yamato to go out and collect samples from the continent. After analyzing some of the plant samples from the continent, XO Shiro Sanada discovers that the floating continent is artificially made by the Gamilans. Meanwhile Raleta, commander of the Gamilas resupply base on the floating continent, gets a message from Gantz, ordering him to dispatch the combat fleet of four ships to attack the Yamato. The Yamato destroys three of the four ships sent after them, and then takes off from the continent. Okita orders Kodai to test fire the Wave Motion Cannon at the floating continent. Kodai fires the gun, destroying the entire continent. Nanbu is thrilled at the power of the weapon, but Okita says that the Yamato 's weaponry is for self-defense only, questioning whether they have done the right thing.

When the Yamato lands on the Saturnian moon of Enceladus to collect some cosmonite and to answer a distress call, Susumu, accompanied by radar operator Yuki Mori and a medical team, investigates the remains of a crashed Earth ship, which he finds to be his brother's vessel, the Yukikaze. Unable to find any clue about Mamoru's whereabouts, they are ambushed by androids sent by the Gamilans and must fight them to escape.

Before leaving the Solar System, the Yamato launches an attack to destroy the Gamilan facilities on Pluto to stop their bombardment of Earth with planet bombs. However, the ship ends up falling for a surprise attack from the enemy's secret weapon, and the crew of Yamato sends their pilot squadron to shut down the Gamilan base.

Having deceived their enemies who believe that the Yamato was destroyed for good, the crew takes the opportunity to stage a counterattack after the Yamato pilots find the location of the main base.

With the Gamilan attacks on Earth stopped, the Yamato prepares to leave the Solar System after giving to the crew members a last chance to contact their relatives that they left behind before resuming their journey to Iscandar.

An operation staged by the Gamilan Leader, Abelt Dessler, forces the Yamato to navigate too close to a star in a desperate attempt to escape, maneuvering around solar flares that can instantly destroy the ship while being pursued by the enemy forces left from the Gamilan base on Pluto.

One of the Gamilan androids retrieved by the Yamato on Enceladus is reassembled and activated in an attempt to learn more about their enemies. Analyzer, the Yamato's own robot, makes contact with the android and manages to befriend it. However, when the android escapes its confinement, the rest of the crew enters a state of alert and searches for it.

The Yamato falls into a dimensional rift and finds itself unable to escape. A Gamilan ship that is also trapped there approaches the Yamato and both crews come to a temporary ceasefire while they work together to escape. After the Yamato escapes the dimensional rift, General Goer meets it with a Gamilan fleet. Instead of fighting the fleet, the Yamato runs away, and the dimensional rift temporarily widens, swallowing the fleet.

Melda Ditz, a Gamilan pilot under the custody of Yamato, claims that their attack on Earth was not unprovoked and that it was the Earthlings who first fired without warning during the first contact between both races years before. Daisuke, whose father was killed in the attack, does not believe her at all until Yamazaki, the sole survivor of his father's ship, reveals to him the truth about what really happened that day.

General Domel of the Gamilans is called back from the front lines by Dessler and reassigned to the Milky Way with orders to stop the Yamato. As the Yamato approaches the edge of the galaxy, the crew becomes under attack by an extra-dimensional submarine.

Taking shelter in an asteroid cluster, the crew of the Yamato discovers the enemy's ability to hide their vessel in sub-space and come up with countermeasures against them.

Returning from a reconnaissance mission, Kodai and Yuki find a derelict Yamato with the rest of the crew missing. As the pair look for clues about what happened, they must face an enemy that can use their own memories against them, assisted by an unexpected ally.

General Domel devises a scheme to trap the Yamato in the midst of a massive fleet of Gamilas warships, but he is unaware of what is taking place behind the scenes on his homeworld.

Badly damaged and in need of provisions, the Yamato lands on a lush, fertile planet named Beemela. As a ground team makes a startling discovery, there are doubts if the ship still has a chance to complete its journey in time and the security force attempts a mutiny.

With the aid of an alien device, Kodai, Yuki, and Sanada explore the control center of an abandoned warp gateway created by an ancient civilization and controlled by the Gamilas. In the midst of a deadly undertaking, Sanada unveils a surprise about the Yamato's navigational functions and his close friendship with Kodai's late brother, Mamoru.

Using the newly reactivated warp gateway, fighter pilot Shinohara scouts the intended shortcut to the halfway point at planet Balun. Once there, however, he discovers a vast fleet of over 10,000 Gamilas ships. However, Okita declares that the Yamato must fight its way through the enemy's massive armada if they want to reach the Large Magellanic Cloud in time.

As the Yamato penetrates the Large Magellanic Cloud, Domel, his name now cleared, engages in a battle of wits with Okita as he prepares a special attack force to stop the Earth vessel once and for all.

Domel's task force engages the Yamato in a nebula dubbed the Rainbow Star Cluster. During a fierce battle in which the Yamato is heavily damaged and its Wave Motion Gun disabled, it is infiltrated by Gamilas operatives tasked to kidnap Yurisha of Iscandar.

Yuki is taken to a Gamilas prison colony called Leptopoda. The Yamato is not far behind, its crew unknowingly seeking to resupply and repair. During a recon mission, Kodai and several others are also taken prisoner just before an old acquaintance foments a riot.

Okita's condition takes a turn for the worse as the rest of the crew learns an unpleasant secret about their quest for Iscandar. Meanwhile, Dessler's grudge against the Yamato becomes more personal than ever, and he prepares to welcome the humans to the heart of Gamilas territory with a weapon of instant destruction.

After being almost destroyed by Dessler's own Wave Motion Cannon, Okita orders the Yamato to launch an offensive at the Gamilas capital. However. Dessler escapes to his space fortress, the "Second Baleras" in a ship with Yuri on board. Dessler then drops a part of the fortress on the capital in an attempt to vanquish the Yamato while sacrificing his own people. While the rest of the crew engages it, Kodai departs to rescue Yuki by himself.

With Dessler gone and the Gamilas ceasing their hostilities, the Yamato, at last, reaches Iscandar. However. Queen Starsha is in doubt about providing the assistance she promised upon seeing that the Earthlings used the technology that she entrusted to them as a weapon of mass destruction. After coming up with a decision, Starsha reveals to Kodai the ultimate fate of his brother as the crew of the Yamato prepares for the long journey back to Earth.

Dessler, who is revealed to be still alive, ambushes the Yamato on its way back to Earth and the time comes for their final showdown.

The Yamato is on the last leg of its journey back to Earth. However, Kodai has little to celebrate as Yuki did not survive the wounds she suffered in the last battle. However, witnessing the suffering of his brother, Mamoru's spirit appears next to him to create a miracle.