Download Brett Dempsey Dr. Arneson 25 April 2016 Alfred Hershey: 1969

Brett Dempsey
Dr. Arneson
25 April 2016
Alfred Hershey: 1969 Nobel Laureate
Introduction
Born on December 4, 1908 to Robert Day Hershey and Alma Wilbur Hershey in Owosso,
Michigan, Alfred Hershey grew up attending several different public schools in Owosso and
Lansing. His father worked for an auto-manufacturing firm, and instilled the importance of
hard work in his son from an early age. Hershey graduated from Owosso High School in 1925,
the same year he enrolled at Michigan State University (AAI, 2016). He was always interested
in the sciences, but developed a keen curiosity for the field of bacteriology during his
undergraduate years. After earning his B.S. in Chemistry from Michigan State in 1930, Hershey
entered a doctorate program in Bacteriology at the same university. His doctoral dissertation
was centered on the chemistry of Brucella, a pathogenic genus of bacteria responsible for
Brucellosis or undulant fever. Hershey’s thesis detailed the chemical mechanisms responsible
for the separation of bacterial components (DNA FTB, 2011).
Following his doctorate training, Hershey accepted a position as an instructor in the
Department of Bacteriology and Immunology at the Washington University of Medicine at St.
Louis. While at Washington University, he worked alongside Jacques Jacob Bronfenbrenner,
who is considered by many to be a key innovator in the field of bacteriophage research. From
1936 to 1939, Hershey and Bronfenbrenner published multiple papers on the growth of
bacterial cultures and the lysogenic properties of bacteriophages (Encyclopedia of World
Biography, 2004). Throughout the early 1940s, he performed his own research involving the
immunologic reaction of phages and other factors that influence phage activity. In 1943,
Hershey began a collaboration with Max Delbruck and Salvador Edward Luria regarding similar
areas of phage study. The three scientists went to Nashville and formed what is known as a
“phage group” where they would come together once a year at Cold Spring Harbor to discuss
their work and encourage research on particular strains of bacteriophages (Encyclopedia of
World Biography, 2004).
1945 proved to be a successful year, as Hershey and Luria both discovered that phage
viruses, along with the bacteria they infect, could experience spontaneous mutations. Later
that year, he and Delbruck discovered that differing strains of bacteriophage could exchange
their genetic material within a dually infected bacterial cell. Hershey referred to this process as
“genetic recombination”. In 1950, he moved to Cold Spring Harbor, New York with his assistant
Marth Chase and became a staff scientist in the Carnegie Institution of Washington’s
Department of Genetics. This would set the scene for his renowned “Hershey-Chase
Experiment” (Encyclopedia Britannica, 2016).
The Awarding of the Nobel Prize
The Hershey-Chase experiment, widely known as the “blender experiment”, was
conducted in 1952 and gave sufficient evidence to allow Hershey to conclude that DNA, rather
than protein, is the genetic material of life. This established the framework for modern
molecular genetics, earning Alfred Hershey a share of the 1969 Nobel Prize in Medicine. Thanks
to this discovery, scientists now knew that DNA is the blue print of all lifeforms on the planet.
This catapulted all areas of biological research, opening a myriad of possibilities for scientists to
investigate. Hershey not only advanced the field of molecular genetics with this discovery, he
uncovered a large piece of the foundation of modern molecular and cellular biology. With how
crucial this finding proved to be, one would have a difficult time devising reasons as to why
Hershey didn’t deserve a Nobel Prize (Berkeley, 2016).
Specifically, Hershey and Chase worked with a DNA virus called T2, a bacteriophage that
infects E. coli. Bacteriophages that are produced within bacteria growing in radioactive
mediums will themselves be radioactive. If radioactive sulfur (35S) is present, they will be
included in the protein coats of the bacteriophage; if radioactive phosphorus (32P) is present,
then the DNA will become radioactive, not the protein. Hershey found that when
bacteriophages containing radioactive phosphorus were allowed to infect nonradioactive
bacteria, all of the infected cells also exhibited radioactivity. Additionally, he found that much
of this radioactivity was passed on to the offspring in the next generation of bacteriophages.
When the bacteria were infected by bacteriophages containing radioactive sulfur on the other
hand, little to no radioactivity could be detected in the infected cells. The radioactive sulfur
detection followed the removal of the virus coats by an electric blender (Kimball, 2014).
These experiments made it clear that the DNA component in the bacteriophages was
injected into the bacterial cells while the protein component remained outside. Furthermore,
they allowed Hershey to conclude that the injected DNA component is the only component that
has the capacity to direct the formation of new virus particles that include protein coats. This
provided even more evidence that DNA is the genetic material. Hershey’s experiments built on
Avery’s 1945 demonstration that genes were DNA, providing further support of this claim.
Many argue that without his work, Watson and Crick would not have been able to determine
the molecular structure of DNA, which earned them a Nobel Prize as well. Hershey’s approach
was so successful due to the fact that he was willing to stray from the classic visual approach
using microscopes. He and his colleagues were true pioneers in the field, focusing on the
molecular level and working with bacteriophages invisible to the naked eye (Kimball, 2014).
Hershey had to overcome the fact that no one in the scientific community knew what
molecule in the cell actually passes on genetic material from parent to offspring. It was
hypothesized that proteins accomplished this task, based on the fact that they are more
complex molecules than DNA. Hershey ardently attacked this question without bias, looking for
evidence regarding the genetic material of life. His discovery transformed the way we ask
questions in biological research, and changed the field of molecular biology forever. Now that
scientists knew that DNA was the genetic material in the body, they could investigate disease
transmission and treatment at many different angles that they once thought not possible.
Referring to Hershey, Luria, and Delbruck (who shared the 1969 Nobel Prize) in her book
“Scientific Elite: Nobel Laureates in the United States”, Dr. Harriet Zuckerman asserted that,
''before the prize finally came to the three founding fathers in 1969, it had gone to 15
molecular biologists and biochemists for investigations built on foundations the three pioneers
had laid down'' (Altman, 1997).
Post-Nobel Prize Activity
Retiring from active research in 1972, Hershey performed most of his post- “blender
experiment” research before winning the Nobel Prize in 1969. In the 17 years between the
Hershey-Chase experiment and the awarding of the Nobel Prize, Hershey continued his work on
phage recombination and genetics. Although he published a few works in these years, none
received the notoriety of the groundbreaking 1952 experiment. In 1962, he became the
Director of the Genetics Research Unit of the Carnegie Institution at Cold Springs Harbor, where
he stayed in this position for three years until his retirement. After 1969, Hershey received no
other notable scientific awards, however his career included memberships in the prestigious
National Academy of the Sciences and the American Academy of Arts and Sciences, an Albert
Lasker Basic Medical Research Award, and the Kimber Genetics Award. Although he retired
from active research in 1974, Hershey frequently visited his former research laboratory and
provided guidance and assistance to those working there. During his free time, Hershey
enjoyed reading, planting trees, and spending time in a woodworking shop. After his
retirement, he pursued other interests including music and computers (Altman, 1997).
Primary Paper Analysis
Independent Functions of Viral Protein and Nucleic Acid in Growth of Bacteriophage
Hershey, A. D. and Chase, Martha.
Journal of General Physiology, Volume 36, Issue 1, 39-56. 1952
“Independent Functions of Viral Protein and Nucleic Acid in Growth of Bacteriophage” is
without argument the most instrumental work in the career of Alfred Hershey. This paper
singlehandedly won him a share of the 1969 Nobel Prize in Physiology or Medicine. It is the
direct result of the notorious 1952 Hershey-Chase or “blender” experiment. The findings
detailed in this paper are considered by many as some of the most crucial scientific discoveries
in the last 100 years (Altman, 1997). This piece of primary literature details the methods
employed by Hershey and Chase to examine the role of DNA and protein in molecular genetics
and bacteriophage growth.
Hershey bases the investigation that is detailed in this paper on previous work by
Doermann, Dissoway, and Anderson. These researchers have shown that bacteriophages T2,
T3, and T4 multiply in a non-infective form. Hershey takes this further by examining the
molecular mechanism involved in T2 growth. He asks whether DNA and protein have different
roles in the growth in bacteriophage T2. Specifically, Hershey uses a variety of bacteriophage
T2 called T2H. “H” refers to the host range mutant of T2, with UV-phage meaning a phage that
is irradiated with ultraviolet light from a germicidal lamp. He also used Esherichia coli as a
bacteria model with varying sensitivities to T2 strains. Resistant bacteria B/2 is resistant to T2
but sensitive to its H mutant, while resistant bacteria B/2h is resistant to both. These particular
bacteria will not adsorb the phages to which they are resistant. The adsorption of either
isotope 35S or 32P to bacteria was measured by mixing the sample in adsorption medium with
bacteria from 18-hour broth cultures that were previously heated. The mixtures were warmed,
diluted with water, and centrifuged. Various assays were then performed with sediment and
supernatant fractions of the mixtures.
The radioactivity of cells was measured using an end-window Geiger counter and dried
samples in order to prevent losses by self-adsorption. For standards regarding the radioactivity
of 32P and 35S, reference solutions were obtained from the National Bureau of Standards.
Additionally, glycerol-lactate medium was chosen over other mediums due to its ability to
permit bacterial growth without causing undesirable pH changes at the low concentrations of
phosphorus and sulfur that were employed. Hershey acknowledges that the radiochemical
purity of the prepared samples is somewhat uncertain. Some might see this as a weakness of
the study, but I would argue that it is a strength. He acknowledges where the uncertainty lies
early in the paper, and proposes that this could be due to the presence of inactive phage
particles and empty phage membranes.
A critical aspect of the paper is the implementation of 35S and 32P.
35S
was used because
it was known that they would be incorporated into the protein coats of the bacteriophages and
therefore will be detectable by end window Geiger counter analysis. It is incorporated into
protein because two of the amino acids, cysteine and methionine, contain sulfur. There are no
sulfur atoms in DNA, so this allows Hershey to differentiate the two molecules.
32P
was used
because it will be taken up by the DNA because of its many phosphorous atoms, causing it to
become radioactive. Additionally, radioactive phosphorous will not be incorporated into the
protein, further allowing Hershey to differentiate between the two molecules.
By employing the method described above regarding the absorption of radioactive
isotopes, Hershey analyzed the sensitization of Phage DNA to DNase by adsorption to bacteria.
The main purpose of this area of investigation was to provide further insight into the molecular
mechanism by which the virus multiplies, including the alteration or removal of the proactive
protein coats of the particles. Hershey derived three main conclusions from this analysis:
1. phage DNA becomes highly sensitive to DNase after adsorption to heat-killed bacteria,
2. DNA of phage adsorbed to live bacteria becomes highly sensitive to DNase after adsorption,
and 3. the DNA of phage adsorbed to unheated bacteria is resistant to DNase. These results
suggest that the DNA of phage adsorbed to live bacteria is protected by cellular structures
immune to DNase activity, most likely a protein coat. Further experimentation confirmed this
claim through microscopy after cycles of freezing and thawing of the DNA.
Hershey also analyzed the respective levels of survival of infected bacteria after removal
of radioactive sulfur and radioactive phosphorus during agitation in a Waring blender. He
found that removal of radioactive phosphorus resulted in an approximately 30% survival rate
while removal of radioactive sulfur yielded an 80% survival rate. Since 32P is incorporated into
DNA and 35S is incorporated into protein, these results provide evidence that the DNA is more
important for cell survival than the protein. This provides support for the claim that DNA
provides molecular instructions for the rest of the cell and is the “blueprint”. Further
investigation with assays of the extracellular isotopes from suspension with agitation and no
agitation illustrated that the bulk of the phage sulfur remains at the cell surface during
infection, and takes no part in the multiplication of intracellular phage. However, he found that
the bulk of the phage DNA enters the cell quickly after adsorption of phage to bacteria. This
provides further evidence that DNA is the genetic material, since it seems to be so much more
involved in the phage multiplication process.
In order to determine whether the DNA or protein is passed from parental phage to
their progeny, Hershey employed a glycerol-lactate medium growth process followed by
sedimentation analysis using centrifugation in order to determine the levels of 35S or 32P in the
supernatant or pellet. He found that in the radioactive sulfur preparation, the radioactivity was
found in the supernatant in the parental phage and little to none in the progeny, suggesting
that the protein was not passed from parent to offspring. In fact, Hershey found that less than
1% of the phage progeny contained the parental radioactivity. On the other hand, he
concluded that 32P was found in the phage progeny at a high rate and was found in the pellet of
the parental phage preparation. This is a groundbreaking finding due to the fact that it suggests
that DNA is in fact the viral genetic material, not protein as previously thought.
The main weakness of this paper is the vague nature by which Hershey describes his
assays. Frequently, he mentions that an assay was used, but never returns to this or provides
details as to what assay was employed. This could be due to the fact that this paper was
written almost 70 years ago and there probably wasn’t very many different types of assays used
in science, however I still think it would improve the paper if he were to go into more depth
regarding the exact methods used.
A strength of this paper is Hershey’s clear portrayal of the results. It was relatively easy
to analyze his results because of the way that he summarized them in numbered lists. This
made the main conclusions from each portion of the experiment and each figure very clear and
easy to recognize. Additionally, Hershey does a great job of identifying where error and
contamination might lie in the experiment and where this could have skewed the results. This
adds to the validity of the experiment and makes it easier to accept the methods that he used.
Summary
Alfred Hershey, a hard-working man from Owosso, Michigan, proved to be a giant in the
field of molecular biology. His discovery provided countless avenues of investigation for
researchers for years to come. Although he began his career in research focused on
bacteriology, his research has been applied across many species including humans. The
Hershey-Chase blender experiment proved to be the most important work of his career, and is
the main reason that Alfred Hershey received a share of the 1969 Nobel Prize in Medicine or
Physiology. “Independent Functions of Viral Protein and Nucleic Acid in Growth of
Bacteriophage” is considered to be the most instrumental publication for his receiving the prize
(Altman, 1997). Hershey completed a variety of valuable research throughout his career, but
none came close to the grandiosity of the blender experiment. Alfred Hershey passed away in
1997, but will always be remembered for his contributions to our understanding of molecular
biology and genetics. “I can only point out a curious fact. Year after year, the Nobel Awards
bring a moment of happiness not only to the recipients, not only to colleagues and friends of
the recipients, but even to strangers”-Alfred Hershey (FamousPeople, 2016).
References
Authors
Title
Source
Altman, Lawrence K.
Alfred D. Hershey, Nobel
The New York Times. 1997.
Laureate for DNA Work,
Dies at 88
The American Association of
Alfred Day Hershey
Immunologists
DNA from the Beginning
AAI: Nobel Laureates.2016.
1-3.
Alfred Day Hershey
DNA from the Beginning.
(1908-1997)
2011.
The Editors of the Encyclopedia
A.D. Hershey: American
Encyclopedia Britannica.
Britannica
Biologist
2016.
Encyclopedia of World Biography
Alfred Day Hershey
Encyclopedia of World
Biography. 2004.
Hershey, A. D., and Chase, Martha
Independent Functions of
Journal of General
Viral Protein and Nucleic
Physiology. 1952. 36,1,
Acid in Growth of
39-56.
Bacteriophage
Kimball, John W.
The Famous People
The Hershey and Chase
Kimball Biology. 19 April
Experiment
2014.
Alfred Day Hershey
TheFamousPeople.com.
2016.
University of California Berkeley
The Hershey-Chase Blender University of California
Experiment
Berkeley Biology. 2016.