The Real Madrid forward was taken down in the box by Nacho as he attempted to find some space to get a shot away, and the referee pointed to the spot.
Ronaldo had given Portugal a dream start winning and scoring a fourth minute penalty. While keeper David de Gea was well positioned to make the save, he couldn't get a strong enough hand on the ball, and had to watch as it continued past him into the goal.
Jose Mourinho lauded Cristiano Ronaldo as a player for "special matches" after his heroics helped Portugal draw 3-3 with Spain on Friday. I am happy with the way the team reacted, we deserved more but football is like that, this can happen to you.
With one point each, Spain and Portugal share the second and third places in their Group B. Iran is leading with three points.More news: GOP scrambles to understand if Trump sunk immigration bill
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And for good measure, he recorded the 51st hat-trick of his career with two further goals, the latter a stunning free-kick - his first successful effort in 45 attempts in a major worldwide tournament - that gave David de Gea no chance.
Cristiano Ronaldo's performance in the club matches have been top notch and he has earned a lot of accolades while playing for Manchester United and Real Madrid. He fooled de Gea by delivering the ball to the top right with authority.
Ronaldo scored his second when David De Gea spilled an easy effort on goal but he saved his last until the final few minutes of the game when he curled home an exquisite free-kick.
However, it was Diego Costa who would level the scores on 24, courtesy of a superb solo goal which saw the ex-Chelsea man twist and turn the Portuguese defence before firing past Rui Patricio. The striker beat defender Pepe to the ball and used several neat moves to clear defenders before firing a low shot into the corner from inside the area. The general consensus is that it was a penalty but Ronaldo went down far too quite easily.
Portugal were starting to look leggy now, as Spain looked set to secure what looked like a deserved victory.