Fiona Hill said the president had instead listened to the views of his personal lawyer, Rudy Giuliani.
Ms Hill called the claims about Ukraine a “fictional narrative”.
The inquiry is assessing if Mr Trump withheld aid to pressure Ukraine to investigate a political rival.
It is illegal in the US to seek foreign help to gain electoral advantage. Mr Trump denies any wrongdoing.
According to a discredited theory, it was Ukrainians or individuals with Ukrainian connections who had interfered in the 2016 vote, rather than Russia.
In a phone call with the Ukrainian president, President Trump urged him to look into the claims as well as open an investigation into Joe Biden, one of the main Democratic presidential candidates.
Thursday was the fifth and last scheduled day of public hearings by the House Intelligence Committee.
Concluding the proceedings, inquiry chairman Democrat Adam Schiff said Mr Trump’s conduct had been far more serious than anything done by President Richard Nixon during the Watergate scandal in the 1970s.
“What we’ve seen here is far more serious than a third-rate burglary of the Democratic headquarters,” he said.
“What we’re talking about here is the withholding of recognition in that White House meeting [and] the withholding of military aid to an ally at war.”
Ms Hill – the former top Russia expert to the White House – accused other members of Mr Trump’s Republican party of sowing doubt about Russian interference in the 2016 elections.
“Based on questions and statements I have heard, some of you on this committee appear to believe that Russia and its security services did not conduct a campaign against our country – and that perhaps, somehow, for some reason, Ukraine did,” she said.
She urged lawmakers not to promote “politically driven falsehoods” that cast doubt on Russia’s interference in US elections.
“This is a fictional narrative that has been perpetrated and propagated by the Russian security services themselves,” she said.
Democratic lawyer Daniel Goldman asked her: “So is it your understanding then that President Trump disregarded the advice of his senior officials about this theory and instead listened to Rudy Giuliani’s views?”
“That appears to be the case, yes,” she replied.
She also warned that Mr Giuliani had been making “explosive” and “incendiary” claims about Ukraine.
“He was clearly pushing forward issues and ideas that would, you know, probably come back to haunt us,” she testified. “I think that’s where we are today.”
She testified to having had a couple of “testy encounters” with Gordon Sondland – the US ambassador to the EU who testified on Wednesday – over Ukraine, because Mr Sondland had not keep her informed of “all the meetings he was having”.
But she later realised he had had a “different remit”.
“He was running a personal domestic political errand, we were concerned with national security,” she said.
US diplomat to Ukraine David Holmes, who also testified at Thursday’s hearing, said his work at the embassy in Kyiv had been overshadowed this year by the actions of Mr Giuliani.
“I became aware that Mr Giuliani, a private lawyer, was taking a direct role in Ukrainian diplomacy,” Mr Holmes said.
Mr Holmes was “shocked” on 18 July when an official from the White House Office of Management and Budget (OMB) announced that military aid to Ukraine was being withheld.
“The official said the order had come from the president and had been conveyed to OMB by Mr Mulvaney with no further explanation,” Mr Holmes said, referring to acting White House Chief of Staff Mick Mulvaney.
Mr Holmes said he had overheard a phone call on 26 July between Mr Trump and Mr Sondhand that was potentially damaging for Mr Trump.
Mr Sondland, in his hearing appearance on Wednesday, confirmed the two men had discussed “the investigations” the president wanted Ukraine to launch during this call.
“When the president came on, [Sondland] sort of winced and held the phone away from his ear like this,” Mr Holmes said, gesturing with his hands.