Group B leaders Italy have a game in hand and a three-point advantage over second-placed Bulgaria, meaning victory in Prague will give Cesare Prandelli's men a six-point cushion at the top.
That outcome would leave Czech Republic eight points shy of top spot but a first win over the Italians since 2002 would see Bulgaria pushed down into third ahead of a September trip to Italy.
The different ramifications of either result are not lost on Juventus defender Bonucci, who is determined to ensure Italy take a "very important step" towards Brazil on Friday night.
Bonucci told figc.it: "We've talked about a lot of things over the last few days but it's the game against the Czech Republic that matters. It's a crossroads in terms of our qualification for the World Cup.
"Winning would take a chunk of points from our opponents and would let us go into next season with a big advantage and would put us a step away from qualifying. The win is crucial for the Czechs, too.
"This is a very important step on our journey to Brazil in 2014."
Both sides arrive at Prague's Generali Arena on the back of wins with Italy having beaten San Marino 4-0 in a friendly last week and the Czech Republic downing Armenia 3-0 in March.
Italy are missing Genoa full-back Luca Antonelli, who limped out of the San Marino game with an ankle injury, while the Czech Republic look to draw on the goalscoring talents of Udinese striker Matej Vydra.
The 21-year-old opened his account for the national team with a brace against Armenia just days after winning the Championship player of the year award during his loan spell with Watford.
Italy last met the Czech Republic at the 2006 World Cup in Germany with the Azzurri - who eventually won the tournament - claiming a 2-0 victory in Hamburg.
The other Group B game sees Armenia host bottom side Malta at the Republican Stadium.
Armenia retain an outside chance of reaching second despite a seven-point disadvantage and Vardan Minasyan's team would match fourth-placed Denmark, who they play next Tuesday, on six points with victory in Yerevan.
Malta are without a point in their bid to qualify but kept the margin of defeat to a single goal for the only time when losing 1-0 to Armenia in Ta' Qali last September.