Holmes was born John Curtis Estes on August 8, 1944, in the small rural town of Ashville, Ohio, about 25 miles (40 km) south of Columbus. He was the youngest of four children born to 26-year-old Mary June (Barton) Holmes, but the father's name, Carl Estes, a railroad worker, is left blank on his birth certificate. Mary Barton had married Edgar Harvey Holmes, who was the father of her three older children - Dale, Edward and Anne. She and Holmes were married and divorced three times, as is documented by wedding certificates dated April 13, 1936, August 13, 1945, and September 12, 1947. At the time of their first marriage in 1936, Edgar Harvey Holmes was 35 years old and divorced, while Mary Barton was 17. After divorcing for the third and final time, Edgar and Mary each married one more time.

When Holmes was age seven, his mother married Harold Bowman on December 31, 1951. Shortly afterward, Holmes and his family moved from Columbus and settled in the small town of Pataskala, Ohio, about 10 miles east of Columbus. Holmes recalled that Bowman was a good father until Holmes' younger half-brother David was born, at which point Bowman reportedly lost interest in his stepchildren and began neglecting them.

In April 1965, Holmes found work as a forklift driver at a meatpacking warehouse in nearby Cudahy, California. However, repeated exposure to the freezing air in the large walk-in freezer after being outside inhaling the desert-hot air caused him severe health problems, leading to a pneumothorax of his right lung on three separate occasions during the two years he worked there.

Another controversy regarding whether Holmes ever achieved a full erection, although much of his early work clearly revealed he was able to achieve a substantial erection. A popular joke in the 1970s porn industry held that Holmes was incapable of achieving a full erection because the blood flow from his head into his penis would cause him to pass out. Fellow film Actress Annette Haven stated that his penis was never particularly hard during intercourse, likening it to "doing it with a big, soft kind-of loofah".

In 1971, Holmes' career began to take off with a porn series built around a private investigator named Johnny Wadd, written and directed by Bob Chinn. The success of the film Johnny Wadd created an immediate demand for more Johnny Wadd films, so Chinn followed up the same year with Flesh of the Lotus. Most of the subsequent Johnny Wadd films were written and directed by Chinn and produced by the Los Angeles-based company Freeway Films.

With the success of Deep Throat (1972), Behind the Green Door (1972) and The Devil in Miss Jones (1973), porn became chic, although its legality was still hotly contested. Holmes was arrested during this time for pimping and pandering, but he avoided prison time by reputedly becoming an informant for the LAPD. Holmes' "handler" during his time as an informant was LAPD vice detective Thomas Blake. Said Blake of his involvement with Holmes, "it was a pleasure working for him".

Determining the number of films Holmes made during the early part of his career is difficult, because the ad copy rarely named him. Copy that did usually used entirely inconsistent names. For Example, one early "Swedish Erotica" brochure from 1973 has five loops featuring Holmes, each citing a different name.

In 1976, he met 15-year-old Dawn Schiller, whom he groomed and abused. After Holmes became desperate for money, he forced her into prostitution and often beat her, which he did at least once in public.

By 1978, Holmes was alleged to be earning as much as $3,000 per day as a porn performer. Around this time, his consumption of cocaine and freebasing were becoming an increasingly serious Problem. Professionally, it affected his ability to maintain an erection, as is apparent from his flaccid performance in Insatiable (1980). To support himself and his drug habit, Holmes ventured into crime, selling drugs for gangs, prostituting himself to both men and women, as well as committing credit card fraud and various acts of petty theft.

In 1979, along with his younger half-brother David Bowman, Holmes opened in Los Angeles a locksmith shop managed by Bowman and an attached used goods store called The Just Looking Emporium, named by Sharon Gebinini and managed by Dawn Schiller. However, because of Holmes' escalating drug addiction, which distracted him from buying inventory for the Emporium and siphoned its working capital, the Emporium "close[d] its doors forever by the end of September 1980". According to Schiller, "David [kept] his part of the Business open while John remove[d] our inventory and [sold] it all for coke".

In late 1980, a mutual friend introduced Holmes to Chris Coxx, who owned the Odyssey nightclub. In turn, Coxx introduced Holmes to Eddie Nash, a drug dealer who owned multiple nightclubs, including the Starwood Nightclub in West Hollywood. At the same time, Holmes was closely associated with the Wonderland Gang, a group of heroin-addicted cocaine dealers, so called for the location of their hangout: a rowhouse located on Wonderland Avenue in the wooded Laurel Canyon neighborhood of Los Angeles. Holmes frequently sold drugs for the gang.

Holmes was questioned regarding the murders in July 1981, but he was released due to lack of evidence. He refused to cooperate with the investigation and, after spending nearly five months on the run with Dawn Schiller, Holmes was arrested in Florida on December 4, 1981, by former LAPD homicide detectives Tom Lange, who later gained fame for his participation in the investigation of the O. J. Simpson murder case, and Frank Tomlinson, and extradited to Los Angeles. In March 1982, Holmes was charged with personally committing all four murders. After a three-week publicized trial, Holmes was acquitted on June 26, 1982, of all charges except contempt of court. The Holmes Murder Trial was a landmark in American jurisprudence, as it was the first murder trial in America where videotape was introduced as evidence.

Later, after his 1982 murder trial and acquittal, Holmes began a Business partnership with his friend, manager and associate Bill Amerson. They founded and operated a production company named Penguin Productions, where Holmes could be a triple-threat: writing, directing and performing. Holmes appeared in seven of Penguin's 20 productions between 1985 and 1988. After requesting permission to use the name Johnny Wadd from his old Director and friend Bob Chinn, Holmes reprised the detective role for the Penguin Production:The Return of Johnny Wadd (1986) – one of his last films.

During the summer of 1986, Holmes was offered a lucrative deal from Paradise Visuals, which was unaware he was HIV-positive, to travel to Italy to film (what were to be his last) two pornographic films. Holmes' penultimate film was The Rise of the Roman Empress (originally released in Italy as Carne bollente) for Director Riccardo Schicchi. The film starred Holmes, the later Italian Parliament member Ilona "Cicciolina" Staller, Tracey Adams, Christoph Clark and Amber Lynn. His final film was The Devil In Mr. Holmes, starring Tracey Adams, Amber Lynn, Karin Schubert and Marina Hedman. These last films created a furor when it was revealed later that Holmes had consciously chosen not to reveal his HIV status to his co-stars before engaging in unprotected sex for the filming. Not wanting to reveal the true nature of his failing health, Holmes claimed to the press that he was suffering from colon cancer.

Holmes married Laurie Rose on January 23, 1987, in Las Vegas, after confiding to her that he had AIDS.

During the last five months of his life, Holmes remained in the VA hospital on Sepulveda Boulevard in Los Angeles. He died from AIDS-related complications (according to his death certificate, cardiorespiratory arrest and encephalitis due to AIDS, associated with lymphadenopathy and esophageal candidiasis) on March 13, 1988, aged 43. His body was cremated, and his widow Laurie and mother Mary scattered his ashes at sea, off the coast of Oxnard, California.

After Holmes' death, the length of his penis continued to be used to market Holmes-related material. For Example, at the premiere of the film Wonderland (2003), patrons were given 13​⁄₂-inch rulers as gag gifts.

Productions in the ‘’Johnny Wadd’’ series: