Du Plessis was booed by the crowd as he walked out to bat, following a ball-tampering incident for which he was found guilty and fined, but cleared to play in the match.
But he was his team's lone hope with a determined knock after the loss of seven wickets had the tourists struggling against Steve Smith's revamped team.
At the dinner break ahead of the much-anticipated final evening session with the pink ball under lights, the Proteas were 165 for 7 with du Plessis on 65 and Kyle Abbott one.
John Hazlewood was proving the destroyer with four wickets for 36.
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Dashing wicketkeeper Quinton de Kock, who has scores of 84, 64 and 104 in the series, had a failure by comparison, managing just 24 off 33 balls.
De Kock again won his battle with Australia's spinner Nathan Lyon, but perished when he edged Hazlewood to wicketkeeper Matt Wade.
Vernon Philander lost a third umpire review when 'Snicko' detected the faintest of edges off Hazlewood to Wade for four nearing the dinner break.
Opener Steve Cook had an early reprieve to go on to score 40 before he fell to Mitchell Starc.
Cook had a major let-off on four, when he was struck on the back pad by Starc and given out leg before wicket in the day's third over.
But he was called back to the crease when replays picked up a front-foot no-ball bowled by Starc.
But Cook fell in the fifth over of the middle session when he sparred at Starc and nicked to Smith at second slip.
Starc recovered from his early Cook blunder to entice a thick edge from Dean Elgar to Usman Khawaja at third slip for five.
He received ironic cheers from the home crowd when replays confirmed it was a legal delivery.
English-born debutant Matt Renshaw then took his first Test catch to dismiss Hashim Amla for five off Hazlewood.
The 20-year-old opener snapped up an ankle-high catch at first slip to claim the big scalp of Amla.
Hazlewood struck again in the over after drinks, coaxing an inside edge off JP Duminy to give Wade a catch for five.