Over a week on from the 20mph speed limit and deputy climate change minister Lee Waters, who headed the introduction of the law change, will face a Senedd vote of no confidence today.
As of September 17, 30mph roads have been reduced to 20mph where cars mix with pedestrians and cyclists.
The move has proved controversial with a poll for ITV Wales revealing that just 33% of people support the policy, while 61% are opposed to it.
A Senedd petition calling for the reversal of new speed limits on most built-up roads has since become the most signed petition in the history of the Welsh Parliament.
The number of signatures have risen sharply, with around 433,000 people having put their names to it.
The Welsh Conservatives said Mr Waters' move will cost the Welsh economy up to £9 billion pounds.
A spokesperson for the party said the new limit is "slowing emergency services down" and "damaging livelihoods and jobs".
They continued: "The Welsh Conservatives are bringing forward a vote of no confidence in the Deputy Minister for Climate Change given the record-breaking number of signatories to the petition.
"It’s clear, the people of Wales do not want blanket 20mph speed limits. They don’t want the Labour Minister’s anti-motorist agenda. His position is not tenable."
Commenting ahead of the vote of no confidence, Welsh Conservative Shadow Transport Minister, Natasha Asghar MS said: “Labour’s Deputy Minister is attempting to undermine the will of the overwhelming majority of Welsh people because he is unable to admit he has got it wrong on his blanket 20mph policy. The Deputy Minister must park his pride and roll back on his anti-road, anti-worker and anti-motorist agenda.
“The people of Wales have had enough. His position is untenable, it’s time for him to go.”
A Welsh Government spokesperson said: “The introduction of a 20mph speed limit in mainly residential areas is designed to save lives and make our communities safer for everyone, including motorists.
"It has been thoroughly researched, voted on in the Senedd and received the backing from a majority of Senedd Members. There has been extensive consultation and it has been trialled in communities across Wales.”