The Israelis were shot dead in the West Bank on Tuesday.
The military wing of the Palestinian Islamist movement, Hamas, said it had carried out the "heroic operation".
The Israeli and Palestinian leaders are to meet on Thursday for the first direct talks in two years.
Ahead of the meeting in Washington, the White House condemned the killing in the "strongest possible terms," adding that all parties must "continue working to achieve a just and lasting peace in the region".
As she went into a meeting with the Israeli Prime Minister Benjamin Netanyahu on Tuesday evening, US Secretary of State Hillary Clinton offered security assurances to Israel.
"We pledge to do all we can always to protect and defend the state of Israel and to provide security to the Israeli people," Mrs Clinton said.
Mr Netanyahu's spokesman, Nir Hefetz, said the Prime Minister had "ordered the security forces to act without diplomatic limitations in order to catch the murderers."
"Terror will not determine Israel's borders or the future of the settlements," Mr Hefetz added said on arrival in the US.
Palestinian Authority Prime Minister Salam Fayyad also condemned the attack, saying it went "against Palestinian interests"
The attack took place near Bani Naim after dark on Highway 60, a busy route used by both Palestinians and Jewish settlers.
The four victims - two men and two women, one of whom was pregnant - were reportedly shot at from a passing vehicle.
Video footage showed a white estate car standing at an angle on the side of the road, its windows shot out and its doors riddled with bullet holes.
Officials said all four victims were from the Beit Haggai settlement.
On its website, the Izz al-Din Qassam Brigades, Hamas's military wing, declared "its full responsibility for the heroic operation in Hebron".
Earlier, Hamas spokesman Sami Abu-Zuhri said the attack had been "a natural response to the crimes of the occupation".
When they meet with Mrs Clinton on Thursday, Palestinian Authority President Mahmoud Abbas and Mr Netanyahu will be in the same room for the first time in almost a year.
In preparation for the talks, President Barack Obama is to meet Mr Netanyahu and Mr Abbas individually on Wednesday. He is also scheduled to meet King Abdullah of Jordan and the Egyptian President Hosni Mubarak.
In order to mark the relaunch of direct talks, Mr Obama will host a working dinner on Wednesday evening with all visiting leaders, Mrs Clinton and UN Envoy Tony Blair.
Before leaving the West Bank, Mr Abbas urged "Israel not to miss this historic opportunity for peace", saying that "if there is only a 1% chance of achieving peace we will strive for it".
Mr Netanyahu meanwhile said he hoped for "a peace based on recognition, security, stability and economic prosperity between the two peoples that will endure for us and our children".
Both sides have agreed to place a one-year time limit on the direct negotiations, but correspondents say prospects of a comprehensive deal are slim, as serious disagreements exist on the core issues.