Lion, the 22-year-old shire horse, had pulled ploughs, coaches and carriages at Beamish Museum in County Durham since 2003.
He was owned by the museum's stockman Jim Elliott.
Hundreds of people have paid tribute on Facebook and a museum spokeswoman said Lion featured in multiple TV shows and films filmed at Beamish.
The spokeswoman said Lion was a "very amenable character" and had a "solid-as-a-rock demeanour".
She said: "Lion could turn his hoof to most things, from ploughing and farm work to pulling omnibuses, charabancs, road sweepers and delivering goods on site. All being done with incredible ease, he also gave a lot of people a lot of pleasure, whether doing traditional experiences or greeting visitors.
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"Lion was a firm favourite with visitors, staff and volunteers alike and will be a huge miss."
The museum said Lion won multiple agricultural competitions as part of Mr Elliot's ploughing team alongside fellow shire horse Prince.
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