The Women’s Tennis Association said Novotna, who had cancer, “died peacefully, surrounded by her family”.
The Czech player had lost in the Wimbledon final in 1993 and 1997 before winning the Grand Slam tournament when she beat Nathalie Tauziat in 1998.
Novotna who was 49, captured the hearts of fans when she burst into tears after losing to German great Steffi Graf in 1993 and was consoled by the Duchess of Kent.
“Jana was an inspiration both on and off court to anyone who had the opportunity to know her,” said WTA chief executive Steve Simon.
“Her star will always shine brightly in the history of the WTA. Our condolences and our thoughts are with Jana’s family.”Novotna was renowned for her serve and volley game and achieved a career-high singles ranking of number two.
In addition to her only singles Grand Slam win at Wimbledon, she claimed 12 Grand Slam doubles titles and four in mixed doubles.
She was also inducted into the Tennis Hall of Fame in 2005.