The Weighted Companion Cube from Portal is one of the game's most famous icons and is prominent in the game's merchandise, as well as a popular subject in fan art. The companion cube does not speak or move or threaten to stab you and only appears in a single level of the game, but is much more well known than the protagonist Chell. Hell, he even got a trope named after him.
The defective personality cores in the last act of Portal 2.
"Y'know what I hope is in space? Fire. I hope you go to space, and you catch on fire."
"The probability of you dying violently in the next 5 minutes is 87.61%."
It's also probably worth noting that the only reason there are so few other voice actors in the game is because they blew an inordinate amount of their budget to get him for that one role.
On a similar note, Liam Neeson lends his voice talent to the main character's father in Fallout 3. Although most of the main quest centers around your father, he only appears in the very beginning of the game and for a brief period in the third act. Tragic, really.
Also, Malcolm McDowell's performance as President Eden is brief but brilliant, and it wouldn't be Fallout without Ron Perlman's intro: "War. War never changes."
Harold, the mutant from the previous two games who had a plant sprouting out of his head by the second, makes an appearance for a sidequest in the third, the plant on his head turning into a full-blown tree with Harold stuck inside.
Sergeant Dornan in Fallout 2, one of the most memorable NPC's despite being little more than an extra. Despite his relative insignificance to the plot (you can rather easily bypass him at camp Navarro), the Sarge's got a "talking head" and voice acting which must be heard to be believed.
Peter Jessup voices Sovereign for two scenes, yet manages to freak out the entire fan community with a single, all-encompassing Badass Boast on behalf of the Reapers;
Sovereign: You exist because we allow it. And you will end because we demand it.
" I am Sovereign. And this station is mine!"
Matriarch Benezia, Liara's mother and Saren's right-hand woman, only actually shows up for two scenes and one audio recording before she's killed whilst performing Suicide by Cop, as it's the only way to free her from indoctrination.
Matriarch Aethyta, Liara's father. She affects one small sidequest, and has only one set of conversation options, which show that she's Seen It All in her thousand-year life and has no qualms about sharing it. The cool old ladies don't get much cooler, or older.
"I saw a krogan drink a liquified turian on a dare six or seven centuries back. Nobody came out of that one looking pretty."
A slightly more traditional example from Mass Effect 2 is Adam Baldwin's brief appearance as Kal'Reegar, a quarian commando with a rocket launcher. Despite only appearing in two places, he's gained a huge fan following.
Kal'Reegar: The geth might get me, but I'm not dying of an infection in the middle of a battle! That's just insulting!
Okeer. Literally a one-scene wonder - a highly intelligent and borderline philosophical character who serves as a traditionalist counterpart to Wrex's reformist idealism, Okeer eloquently explains traditional krogan morality to Shepard while sounding utterly badass. And he provides Shepard with Grunt, sacrificing himself in the process.
Private Jenkins, the doomed squadmember from the first game who dies two minutes into your first mission. Even in-universe, he's occasionally mentioned fondly by various Normandy crewmembers.
Niftu Cal, the drugged up volus who think's he's a biotic god.
Rila, Samara's middle daughter. She only appears at the end of the monastery mission, but her Heroic Willpower and Last Stand really left a mark.
Lancer in Fate/stay night. He is the first enemy encountered in the game's introduction and does little but fight with Archer and all but kill Shirou, after which he spends most of the time in the background . He only gets a bit of spotlight in Unlimited Blade Works when he helps Rin and Shirou by keeping Archer occupied, and later rescues Rin by killing Kotomine and scaring off Shinji while missing his heart. Oh, and inadvertently saving Shirou post-humously by making Gilgamesh sooty. Outside of that he's basically the poster boy for The Worf Effect, being killed off by the The Dragon in the other two routes. Yet he's almost as GAR as Archer for the fandom.
On the other hand, in Fate route...he DOES fight Gilgamesh in a duel to the death Heroic Sacrifice...against orders. And without complaints.
In World of Warcraft there are a very few high-ranking NPCs that get an occasional memorable scripted scene.
In Wrath of the Lich King, when Tirion Fordring appears for any significant speaking role, it's usually worth waiting and watching.
Large Ham or not, the ex-Death Knight, Thassarian, easily steals the spotlight of any questline in which he is involved.
Grand Apothecary Putress. He sends low-level players out on some really shady quests and then you don't hear from him for 40 levels. At level 75, though, he re-emerges at Wrathgate - crashing a war between Horde/Alliance and the Scourge - with a fucking vengeance.
"Did you think we had forgotten? Did you think we had forgiven? Behold, now the terrible vengeance of the Forsaken! Death to the Scourge, and DEATH TO THE LIVING!
And then did that sterling dead man own seventy shades of bejeezus out of the Horde, the Alliance and the Scourge - Including Bolvar 'Dragon Puncher' Fordragon, Saurfang Jr. and the eponymous Lich King, fucking Arthas himself. Many players - particularly Undead players - would have sworn allegiance to that man on the spot.
Although technically a One Level Wonder, Father Grigori in Half-Life 2, who like most examples achieves this with his first line. "You have already met my, hehe, 'congregation'."
Mike the helicopter pilot in Resident Evil 4. Notable for blowing shit up as the only backup you'll ever recieve in the game, and promptly getting shot down as soon as the one level is finished. Players also remember him because Leon shows far more pathos over Mike's death than even the two police officers he traveled with at the beginning of the game.
The bug-like sentry turrets in Doom 3 are only seen in action at one small portion of the game, but their presence gives the player an immense relief. They sport a bright headlight and a machine gun, and will quickly locate and kill any hidden enemies while spouting a series of agitated beeps and clicks. Arguably, they could be the only characters in the game the player may feel an emotional connection to.
The Green Biker Dude from the Mega Man X2 intro. On screen for barely 5 seconds, and he's still one of the most memorable X series characters.
Fire Emblem has so, so many; minor bosses who are only in one stage tend to be among the most beloved in the fanbase. This includes the likes of Batta the Beast, Glass, Denning, Gheb, and Oliver, the last of which was popular enough that he returned in the next game as a recruitable unit. There's also the 3-13 Archer, a nameless allied NPC from Radiant Dawn, who is often considered one of the greatest units in the game. However, one of the most major examples is Dheginsea, the Black Dragon King, who is one of the world's most powerful beings and plays an essential role in the backstory of the Tellius games... he appears three times between the two games.
A minor Carta thug in Orzammar has a performance so over the top when you beat him, you'll want to bring him along for the rest of the game.
The pirate queen Isabela's only major scene happens when you visit Denerim's brothel for the first time, and she'll teach the Duelist specialization for Rogues if you beat her at a rigged game of cards...or if you have sex with her, which can become a three- or foursome depending on your companions. Her character was so popular (mostly due to players wondering how to get the foursome, or have an all-girl threesome), that she was brought back in Dragon Age II as a full party member and possible love interest - as promiscuous, morally suspect, and far more pirate-y than ever, and a bottomless well of Double Entendres and horrible puns.
In Dragon Age: Inquisition, Movran the Under, the Avvar chief who's response to the death of his son is to throw live goats at Skyhold (as in the building itself). He appears only while the player decides his punishment, hamming it up the whole time.
The opening cinematic of StarCraft Brood Wars, we get to see what would be a terran defense against the zerg, as well as some of the units not being affected by Gameplay and Story Segregation and fighting as good as they should in theory (this means that a single marine will lose against a lone zergling, and that the firebats attack can kill both enemy and friend), afterwards we get to see a random marine who easily takes down two zergling, which a previous one couldn't even after depleting his entire ammo with his rocket launchernote In the Starcraft universe, the type of Gauss Rifle that was used in the first war had a secondary fire option, , earning mentions of being the first marauder.
Note that this marine is black, wearing sunglasses, and is more concerned with rocking out to guitar solos than his impending death.
Poseidon in God of War 3. All of the other gods have at least one other appearance in the game, but Poseidon only shows up during Kratos and the Titans' siege of Mount Olympus due to being killed during it. But boy, is his role in that part memorable.
In the CD-i games, several minor characters are immensely popular like Morshu, who only has two scenes.
Morshu: "Mmmmmm... richer."
And of course, King Harkinian. He only appears in the intro to Faces of Evil and is only important to the plot of Wand of Gamelon, and even then only appears in the intro and ending. Come the advent of YouTube Poop, he's one of the most popular characters EVER.
Link in The Wand Of Gamelon He only appears in the opening and closing cutscenes.
Golden Sun fandom is largely convinced that Rief's sister Nowell from Dark Dawn will return as a player character in the next game. All we know about her presently is that she doesn't like her little brother getting into danger, and that she does like Captain Piers enough to go for an unplanned joyride with him.
There's exactly one playable Pandaren in the whole Warcraft series (so far), he's an optional character in a campaign that you don't even need to play in a regular playtrough, he became so popular that Blizzard have been teasing World of Warcraft players with making Pandaren a playable race since the early stages, they will finally get their own expansion (they included the race's name in the title of the expansion). Oh, and the character wasn't even planed to be included in the game, one of the designers made a concept artwork as an april's fool joke.
Kingdom Hearts 3D has one that seemed to exist solely for the trailers, and lasted for all but two seconds. Oh, hi there, Vanitas. Oh, bye, Vanitas!
Nightmare only appeared very briefly as the Final Boss at the very end of one game in the Kirby franchise, yet he remains one of its most iconic villains. Probably helps that he's the one who started the Vile Villain, Saccharine Show trend the series is famous for.
Speaking of which, Zero from Kirby's Dream Land 3 is equally famous, appearing right out of nowhere with absolutely no foreshadowing and immediately becoming the most memorable part of the game, largely for the shocking scene where it rips it's own bleeding eye out.
Drawcia from "Kirby Canvas Curse." She only has one appearance, yet she starts the trend of the True Final Boss soul forms of Big Bad main villains.