Har Gobind Khorana. Photo: nobelprize.org
January 9, 2022, was the 100th birth anniversary of Har Gobind Khorana, a pioneering chemical and synthetic biologist, decipherer of the genetic code and a beloved mentor. His friend Uttam L. RajBhandary, a professor of molecular biology at the Massachusetts Institute of Technology, once said that Khorana believed sincerely in Otto Loewi’s motto: “We must be modest except in our aims” – a statement amply borne out in his life-story.
Khorana was born in 1922, in British India to a poor family that placed a premium on education. In 1945, after receiving his bachelor’s and master’s degrees in chemistry from Punjab University, he received a scholarship from the Ministry of Agriculture to complete a PhD. Initially it was to be on fungicides and insecticides but the need to accommodate veterans from the second World War prompted the Indian High Commission to change his research focus to organic chemistry.
After he completed his PhD, and having saved a portion of his stipend, Khorana funded his own post-doctoral position under Vladimir Prelog. He later recalled that Prelog taught him to see beauty in chemistry, work and effort. Apart from publishing two papers, Khorana learned German to better understand cutting-edge chemical research.
In 1949, he returned to a newly independent and chaotic India, where, in spite of his qualifications, he was unable to find a job. But his connections with the scientific community resulted in a position at the lab of Alexander Todd in the UK. There, he contributed to the synthesis of the building blocks of DNA.
The story of the discovery of heredity is a long one, starting from Gregor Mendel in the 19th century, but many concepts that are common today, such as the double helix structure of DNA, were actually being worked out at Cambridge at the time of Khorana’s stay there.
Imagine DNA is a twisted ladder. Each rung of the ladder is made of one of four possible bases: adenine (A), cytosine (C), thymine (T) or guanine (G). The backbone, or the ladder’s two poles, is made of sugar molecules attached to each other by phosphate bonds. So when you break up DNA, you’ll find that its individual chemical units are sugars, phosphate bonds and bases.
Khorana and his co-workers at the Todd lab created these building blocks through chemical reactions. It was a big step towards unravelling the puzzle of heredity.
Gordon Shrum, the director of the University of British Columbia, invited Khorana to start his laboratory there in 1952. The university didn’t offer many facilities, Shrum believed “organic chemical research was the cheapest to carry out, requiring only test tubes”. With Shrum’s encouragement, Khorana decided to focus on the synthesis of oligonucleotides (short portions of DNA).
This topic was critical to advancing the study of genes and for synthetic biology. This work would also open the way to understanding mRNA – the polymeric molecule at the heart of the Pfizer-BioNTech and Moderna vaccines against COVID-19.
To build on this work, Khorana moved to the University of Wisconsin in 1960, and set out to tackle the defining problem of his era: how does the sequence of bases in DNA specify the sequence of amino acids in proteins? He discussed the challenges he faced on this path in his Nobel Prize lecture (1968), which he began with these words:
“Recent progress in the understanding of the genetic code is the result of the efforts of a large number of workers professing a variety of scientific disciplines. Therefore, I feel it to be appropriate that I attempt a brief review of the main steps in the development of the subject before discussing our own contribution which throughout has been very much a group effort.”
One of the first obstacles was that Khorana’s team needed large quantities of DNA and RNA. Their solution was to use an enzyme called DNA polymerase to speed up the reactions by which oligonucleotides are made. In the process, they discovered fundamental properties of DNA and DNA polymerase that are important to pass genetic information from one generation to the next. They also found that RNA is a necessary intermediary step between the DNA and protein synthesis.
Proteins are made of polypeptides; each polypeptide consists of a chain of amino acids. Over time, Khorana’s team was able to streamline a process to synthesise polypeptides outside the cell using a defined DNA template. After this, they moved on to testing the trinucleotide repeat hypothesis.
According to the hypothesis, based on the work of Marshall Nirenberg, the DNA gives a cell instructions to manufacture amino acids using three bases at a time. A set of three such nucleotides was called a codon.
To test, Khorana’s team synthesised DNA fragments with two alternating bases, such that they had two codons. For example, a fragment with T and C alternating could have two types of codons: TCT and CTC. Through a series of logically progressive experiments, they were able to figure out which codon corresponded to which amino acid.
Ultimately, they were able to put together a map of oligonucleotides and their corresponding amino acid sequences. This work dovetailed seamlessly with the results from Nirenberg’s experiments, proving their validity.
This way, Khorana, Nirenberg, Robert Holley and many other scientists together built the Rosetta stone of biology: the codon table. This is a basic tool that scientists around the world use today to identify, study and manipulate genes.
Khorana continued to pioneer new areas of study throughout his career. After joining the Massachusetts Institute of Technology in 1970, he synthesised the first completely artificial gene and then incorporated it into a living organism – providing the first demonstration of synthetic biology. This proof of principle led to many applications, including engineering of microbes to clean oil spills, gene therapy and synthetic genomes. He also contributed to the then-nascent field of signal transduction and helped to understand how vision works.
In keeping with Loewi’s motto, Khorana was always modest except in his aims. Or, as he put it once, “I always work on big problems.”
Deepika Calidas is a biochemist. Her last position was as a postdoctoral fellow at the Johns Hopkins School of Medicine.