While working at the Medical Clinic of the University of Freiburg, Krebs met Kurt Henseleit, with whom he investigated the chemical process of urea formation. In 1904, two Germans A. Kossel and H. D. Dakin had shown that arginine could be hydrolysed by the enzyme arginase to form ornithine and urea in inorganic reaction. Based on this reaction, Krebs and Henseleit postulated that in living cells, similar reaction could occur, and that ornithine and citrulline could be the intermediate reactions. Krebs started working on the possible method for the synthesis of arginine. Using his Warburg manometer, he mixed a slice of liver with purified ornithine and citrulline. He found that citrulline acted as a catalyst in the metabolic reactions of urea from ammonia and carbon dioxide. He and Henseleit published their discovery in 1932. Thus the ornithine cycle was established, and it was the first metabolic cycle to be discovered.

Krebs was born in Hildesheim, Germany, to Georg Krebs, an ear, nose, and throat surgeon, and Alma Krebs (née Davidson). He was the middle of three children, older sister Elisabeth and younger brother Wolfgang. He attended the famous old Gymnasium Andreanum in his home town. Before completing his secondary school education (by six months) he was conscripted into the Imperial German Army in September 1918, during World War I. He was allowed to appear in an emergency examination for the higher school leaving certificate, which he passed in such a good grade that he suspected the examiners of being "unduly lenient and sympathetic". The war ended after two months and his conscription ended. He decided to follow his father's profession and entered the University of Göttingen in December 1918 to study Medicine. In 1919 he transferred to the University of Freiburg. In 1923 he published his first technical paper on tissue staining technique, the study which he started under the guidance of his Teacher Wilhelm von Mollendorf in 1920. He completed his medical course in December 1923. To obtain a medical license he spent one year at the Third Medical Clinic in the University of Berlin. By then he turned his ambition from becoming a practising physician to medical researcher, particularly towards chemistry. In 1924 he studied at the Department of Chemistry at the Pathological Institute of the Charité Hospital, Berlin, for informal training in chemistry and biochemistry. He finally earned his MD degree in 1925 from the University of Hamburg.

In 1926 Krebs joined Otto Heinrich Warburg as a research assistant at the Kaiser Wilhelm Institute for Biology in Dahlem, Berlin. He was paid a modest 4800 marks per year. After four years in 1930, with 16 publications to his credit, his mentor Warburg urged him to move on and he took up the position of Assistant in the Department of Medicine at the Municipal Hospital in Altona (now part of Hamburg). The next year he moved to the Medical Clinic of the University of Freiburg. At Freiburg he was in-charge of about 40 patients, and was at liberty to do his own research. Before a year was over at Freiburg, he, with a research student Kurt Henseleit, postulated the metabolic pathway for urea formation, now known as the ornithine cycle of urea synthesis. (Sometimes also referred to as the Krebs–Henseleit cycle. Together they also developed a complex solution for studying blood flow in arteries or perfusion ex vivo called Krebs–Henseleit solution or buffer.) In 1932 he worked out the basic chemical reactions of urea cycle, which established his scientific reputation.

Krebs's life as a reputed German scientist came to an abrupt halt because of his Jewish ancestry. With the rise of Hitler's Nazi Party to power, Germany decreed the Law for the Restoration of the Professional Civil Service (the removal of all non-Aryans and anti-Nazis from professional occupations). Krebs received official dismissal from his job in April 1933, and his Service was terminated on 1 July. An admirer, Sir Frederick Gowland Hopkins at the University of Cambridge, immediately came to his rescue and persuaded the university to recruit Krebs to work with him in the Department of Biochemistry. By July 1933 he settled in Cambridge with financial support from the Rockefeller Foundation. Although he was restricted to bring only his personal belongings, he was fortunate to be allowed to take his equipment and research samples to England, as they proved to be pivotal to his later discoveries, especially the manometer developed by Warburg specifically for the measurement of oxygen consumption in thin slices of tissues; it was the basis for his research. He was appointed as Demonstrator in biochemistry in 1934 and in 1935 the University of Sheffield offered him a post of Lecturer in Pharmacology, with a more spacious laboratory and double the salary; he worked there for 19 years. University of Sheffield opened a Department of Biochemistry (now Department of Molecular Biology and Biotechnology) in 1938 and Krebs became its first Head, and eventually Professor in 1945. Krebs took over the running of the Sorby Research Institute in 1943. In 1944, the British Medical Research Council established the MRC Unit for Cell Metabolism Research at Sheffield, and Krebs was appointed as the Director. With this his laboratory became so expanded that the locals jokingly nicknamed it "Krebs's Empire". He moved with his MRC unit to the University of Oxford in 1954 as Whitley Professor of Biochemistry, the post he held till his retirement in 1967. The editorial board of Biochemical Journal extended their good wishes on his retirement, but in return he promised to keep them busy (by producing scientific papers). He continued research and took his MRC unit to the Nuffield Department of Clinical Medicine at the Radcliffe Infirmary, Oxford. From there he published over 100 research papers.

Krebs sent a short manuscript account of the discovery to Nature on 10 June 1937. On 14 June he received a rejection letter from the Editor, saying that the journal had "already sufficient letters to fill correspondence columns for seven or eight weeks", and encouraging Krebs to "submit it for early publication to another periodical." Krebs immediately prepared a longer version titled "The Role of Citric Acid in Intermediate Metabolism in Animal Tissues", which he sent to the Dutch journal Enzymologia after two weeks and was published in two months. It was followed by a series of papers in different journals.

Krebs met Margaret Cicely Fieldhouse (born 1913) when he moved to Sheffield in 1935. They married on 22 March 1938. Krebs later described his life in Sheffield as "19 happy years". They had two sons, Paul (born 1939) and John (born 1945), and a daughter, Helen (born 1942). John (Sir John Krebs, and later Baron Krebs) became a renowned ornithologist, Professor at the University of Oxford, Principal of Jesus College, Oxford, and Member of the British House of Lords.

Krebs became a naturalised British citizen in 1939. He was elected Fellow of Trinity College, Oxford, 1954-1967. He was elected to the Royal Society in 1947. In 1953 he received the Nobel Prize in Physiology or Medicine for his "discovery of the citric acid cycle." (He shared the Nobel Prize with Fritz Lipmann.) For the same reason he was given the Albert Lasker Award for Basic Medical Research in 1953. The Royal Society awarded him its Royal Medal in 1954, and Copley Medal in 1961. In 1958 he received the Gold Medal of the Netherlands Society for Physics, Medical Science and Surgery. He was knighted in 1958 and was elected Honorary Fellow of Girton College, Cambridge University in 1979. He was the Original Member of the Society for General Microbiology, which conferred him Honorary Membership in 1980. He received an honorary doctorate from 21 universities.

Krebs continued to add more details to his citric acid cycle. The discovery of acetyl-CoA in 1947 by Fritz Albert Lipmann was another major contribution. However, this new discovery posed a Problem in his classic reaction. In 1957 he, with Hans Kornberg, found that there were additional crucial enzymes. One was malate synthase, which condenses acetate with glyoxylate to form malate, and the other was isocitrate lyase, which provides glyoxylate for the reaction by cleaving it from isocitrate. These two reactions did not follow the normal citric acid cycle, and hence the pathway was named the glyoxylate bypass of the citric acid cycle, but is now known as the glyoxylate cycle.

After a brief illness, Krebs died on 22 November 1981 in Oxford.

The University of Sheffield has The Krebs Institute, founded in 1988. It is a research centre covering interdisciplinary programmes in biochemical research.

In 1990 the Federation of European Biochemical Societies instituted the Sir Hans Krebs Lecture and Medal, which was endowed by the Lord Rank Centre for Research. It is awarded for outstanding achievements in biochemistry and molecular biology.

The University of Oxford had a building named Hans Krebs Tower, which was occupied by the Department of Biochemistry. In 2008 a new building for the Department of Biochemistry was constructed, on which a plaque was placed on 20 May 2013 by the Association of Jewish Refugees. The plaque was unveiled by John, Lord Krebs, and the inscription reads:

The Biochemical Society offers Krebs Memorial Scholarship to a postgraduate (PhD) student working in biochemistry or an allied biomedical science at any British university. As of 2014, the scholarship is worth £18,500 and is given for a year, but is extendable up to three years.

In May 2017, Krebs's Nobel Prize medal has been auctioned off for £225,000 (around $351,225). The proceeds will be used to found the Sir Hans Krebs Trust which will provide funding for doctoral students in the biomedical field and support chemists who had to flee their home countries.