The plan provides for a joint response between both countries in the event of an incursion or a limited attack from the North, officials say.
Help from the US - which has 28,000 troops in South Korea - has until now been optional in minor skirmishes.
Regional tension remains high after the North's third nuclear test last month.
The US already offers South Korea a "nuclear umbrella", but Cold War experts have pointed out that while nuclear deterrence may address the possibility of all-out war, it does not deter low-level incidents.
Under the new plan, South Korea will be able to call on US assistance should Pyongyang follow through with its recent threats, for example to attack remote South Korean islands, says the BBC's Lucy Williamson in Seoul.
"This allows both nations to jointly respond to the North's local provocations, with the South taking the lead and the US in support," South Korean defence ministry spokesman Kim Min-seok said on Monday.
"It will have the effect of preventing the North from daring to provoke us," he added of the deal, which was signed on Friday.
The "provocative" acts that the plan seeks to address include incursions on the border and by low-flying aircraft, and attacks on border islands.
The new plan was conceived in 2010, after North Korea shelled a border island. A South Korean warship also sank that year, leaving 46 sailors dead. South Korea said North Korea torpedoed the ship, but Pyongyang denied this.
Last month the United Nations imposed fresh sanctions against North Korea following its nuclear test on 12 February.
Pyongyang has responded with escalating rhetoric both to this and US-South Korea joint military drills which it bitterly opposes.
It says it has scrapped the Korean War armistice and ended non-aggression pacts with Seoul.
South Korea says North Korea cannot unilaterally dissolve the armistice and has called on Pyongyang to tone down its language.
