Klitschko told his UDAR party that he plans to run for mayor of Ukraine's capital city, Kiev, paving the way for current presidential favorite, businessman Petro Poroshenko, in the May 25 vote.
The move is likely to ensure that both men cement powerful positions in Ukraine's new government and block the chances of a full return to power from the country's former prime minister, Yulia Tymoshenko.
The May 25 presidential election and Kiev mayoral vote are taking place against the backdrop of Russia's recent annexation of Crimea, Ukraine's dire economic straits and rumblings of discontent in the country's mainly Russian-speaking eastern provinces.
Both Klitschko, 42, and Poroshenko, 48, played prominent roles in the months-long protest movement that led to the toppling of President Viktor Yanukovych in February. The demonstrations were sparked by Yanukovych's decision to back away from closer ties with the European Union and turn toward Russia but grew to encompass widespread discontent with corruption and the lack of democratic freedoms.
"The only way to win is by nominating a single candidate from the democratic ranks," Klitschko said. "This should be a candidate with the greatest support from the people."
Poroshenko, who also owns the popular Channel 5 television station and has served as foreign minister, already leads in the polls for the presidential election and is seen as likely to beat Tymoshenko, who declared this week that she will "be the candidate of Ukrainian unity."
One analyst said Saturday's announcement was a strategic move by Klitschko.
"The alliance of Poroshenko and Klitschko will fundamentally change the configuration of the political field and will cancel out the chances of other candidates," said political analyst Vadim Karasyov. "It is clear that Yulia is not fighting for the post of president, but for a future faction in parliament." As Ukraine moves quickly to build up its governance, it still casts an uneasy eye toward Russia, fearing a possible troop invasion into eastern Ukraine.
President Barack Obama urged Russian President Vladimir Putin to pull his troops back from the border with Ukraine during an hour-long phone call Friday. The Russian leader, who initiated the call, asserted that Ukraine's government is allowing extremists to intimidate civilians with impunity - something Ukraine insists has not happened.
In a televised interview aired Saturday, Russian Foreign Minister Sergey Lavrov expressed concerns over the threat posed by radical nationalists to ethnic Russians in eastern Ukraine, but denied any armed action was imminent.
"We have absolutely no intention or interests in crossing the borders of Ukraine," he said.
Poroshenko announced his candidacy to supporters Friday evening in his childhood hometown of Vinnytsia, holding up a religious icon of the Virgin Mary and child.
Speaking Saturday at the same UDAR congress as Klitschko, Poroshenko said Ukraine needed to unify in the face of aggression, a reference to Russia's annexation of Crimea.
"Our goal is to live in a new way," he said Saturday. "To shape Ukraine in a way that there will be rich, free and honest citizens that will be happy to be Ukrainians and to live in a country respected by the whole world."
Several political party conferences took place Saturday across Kiev ahead of the Sunday deadline for presidential candidates to submit their bids.
Tymoshenko's bid for the presidency was overwhelmingly supported by her Fatherland party, whose representatives have key positions in the government installed after Yanukovych's ouster. The two have been rivals for a decade - she became prime minister after Yanukovych's presidential win in 2004 was thrown out due to vote fraud, and he beat her in the 2010 presidential ballot. She was freed when Yanukovych was ousted after spending two years in jail for charges that many in the West viewed as trumped up by his government.
Addressing supporters on Kiev's historic Sofiivska Square, Tymoshenko heavily condemned both Putin and the annexation of Crimea.
"Putin wanted to punish us Ukrainians for our desire for freedom, for our European choice, for our ability to fight," she said. "If you give me your trust to become president, I will not yield another centimeter of Ukrainian land to the aggressors."
Prime Minister Arseniy Yatsenyuk, a close associate of Tymoshenko, vowed that the elections would properly reflect the will of the people.
"As the head of Ukraine's government, I guarantee that we will hold fair and clean elections," he said.
At its conference, the former ruling Party of Regions voted to expel Yanukovych, who has fled to Russia, along with several other senior officials linked with the now-deposed government. The party nominated Mikhail Dobkin, formerly the governor of the heavily Russian-speaking Kharkiv province, as its candidate for president.
More than a dozen other presidential hopefuls are throwing their hat into the ring. One announced his intentions Saturday at a press conference in Kiev dressed as Star Wars character Darth Vader and flanked by supporters in Imperial Stormtrooper and Chewbacca costumes.
A survey in mid-March by the SOCIS polling organization found Poroshenko leading with a wide margin at 36 percent support among likely voters. Klitschko was seen in second place with 13 percent, Tymoshenko had about 12 percent and Dobkin had over 5 percent support.
In Russian-controlled Crimea, meanwhile, the leadership of the peninsula's Tatars announced plans to hold a referendum on declaring autonomy for their community.
Associated Press writers Yuras Karmanau in Kiev and Laura Mills in Crimea contributed to this report.