The 29-year-old Monegasque father-to-be, who won in Austin two years ago, clocked a best lap in 01:35.291 seconds to outpace the 20-year-old Italian by 0.042 seconds, with four-time champion Max Verstappen of Red Bull 0.189 seconds adrift in third.
Leclerc, whose wife Alexandra Saint Mieux is expecting their first child, had earlier topped opening practice ahead of Verstappen and drove like a man on a mission throughout the day.
Antonelli could clinch the drivers' title to become Formula 1's youngest champion - and Italy's first since Alberto Ascari for Ferrari in 1953 - this weekend if he wins and his main rivals are unable to stay within reach of his points total; teammate George Russell and Ferrari's Lewis Hamilton need to finish in the top eight and top four respectively if Antonelli wins.
Hamilton was fourth for Ferrari, six-tenths off the pace, with Russell fifth ahead of Racing Bulls' Liam Lawson and the McLarens of world champion Lando Norris and Oscar Piastri.
Isack Hadjar was ninth in the second Red Bull and Oliver Bearman 10th for Haas.
Three drivers had grid penalties confirmed between the sessions: five places for Racing Bulls' rookie Arvid Lindblad, ten for Alpine's Franco Colapinto and 20 for Audi's Gabriel Bortoleto, all for exceeding the limits on taking new power unit components.
Verstappen, winner of the last two races, showed he meant business as soon as the pitlane opened for FP2 by accelerating after the two Racing Bulls and, unexpectedly, surging past Lawson at Turn One, much to the New Zealander's surprise.
He swiftly posted a lap in 01:36.581 to go top ahead of Russel, with Antonelli grumbling about his seat 'moving around like crazy', a distraction that required an early stop in the pits.
Norris, on hards, then went top briefly before Russell beat his time but then dropped to second as Verstappen took a tow from team-mate Hadjar to clock 01:35.934 and go clear by a tenth.
Given his need to win, Antonelli's absence for repairs was a setback as his rivals, notably Verstappen and Russell, capitalised before he rejoined the fray - and immediately took third behind them. It was a sign of his pure talent and speed.
By this time, Russell had experienced a curious skirmish with Leclerc which resulted in a black-and-white flag warning him for driving standards. It appeared he had forced the Ferrari driver off track.
After a busy half hour, most switched to soft tyres for some qualifying simulation runs that began with Russell going top before Verstappen and then Leclerc took control, the Ferrari clocking 01:35.291.
To confirm Ferrari's form, Hamilton went third, albeit six-tenths adrift of his team-mate, until demoted a place by the hasty Antonelli grabbing second, just 0.042 off Leclerc's time.