Cheryl Gillan faced criticism on Sunday for selling a home close to the planned High Speed 2 rail route two months before the Government gave the project the go-ahead.
The Wales secretary sold the 17th century terraced house in Amersham, Buckinghamshire, in November for £320,000.
The property is situated about 500 yards from the route of the London to Birmingham section of HS2, which was given the green light last week by transport secretary Justine Greening.
Gillan has previously threatened to resign over HS2, which is bitterly opposed by many of her Chesham and Amersham constituents who complain that it will blight the area.
She said last week that "good progress" had been made in mitigating the effects of the new line with tunnels.
A spokesman for Gillan said her house had been on the market since June 2010 and was sold for 20% under the original asking price.
She sold it because neither she nor her husband were able to use the stairs. Gillan has mobility problems after an illness in 2006 and her husband is 84.
"It had nothing whatsoever to do with HS2," the spokesman said.
But Seb Berry, an independent councillor for the nearby town of Great Missenden, told The Sunday Telegraph: "We now have an MP who sits in the cabinet that has approved a policy that will blight many homes in her own constituency - and yet she no longer owns a property here.
"That will make local people - many whom were in tears over the HS2 decision - very angry."
Shadow Cabinet Office minister Jon Trickett wrote to prime minister David Cameron urging him to investigate whether there had been any breach of the ministerial code, which forbids any conflict between public duty and private interests.
"If the reports in the national newspapers are accurate, it is very likely that Cheryl Gillan has breached the ministerial code," he said.
"If true this is a shocking abuse of her ministerial position and shows complete disregard for the constituents she has abandoned as well as the responsibilities and duties that come with her ministerial post.
"Gillan has cut and run. She has taken an option that simply wasn't available to her constituents and her actions have damaged not just her own reputation and credibility but that of the government."