Some of the most important evolutionary events in Earth’s
history didn’t just create new organisms — they created
new fundamental biochemical processes. And where do
biochemical processes come from? They evolve from other
biochemical processes.

Two of the most important pieces of biochemical innovation
that occurred in the early biosphere — the development of
photosynthesis (which made light energy available to life)
and of nitrogen fixation (which made atmospheric nitrogen
available to life) — may be related to each other because
some of their key enzymes appear to have evolved from a
common ancestor that may be part of a third, significantly
different, biochemical process.

“Photosynthesis was important because it gave life an
enormous energy source and ultimately put oxygen in the
atmosphere,” said Arizona State University biochemist
Robert Blankenship. “Nitrogen fixation — making
atmospheric nitrogen bioavailable — was also a critical
step in the early development of life. We need a good
source of nitrogen for proteins and DNA, but the biggest
source, the molecular nitrogen that we have in the
atmosphere, has a triple bond in it that makes it so
inert that it’s a killer to get at.”

Two new studies, to be presented at the February 2003
NASA Astrobiology Institute General Meeting by researchers
at Arizona State University, provide evidence for the
long-suspected relatedness of the two biochemical
pathways, and find hints of other related pathways that
may be key to understanding the evolutionary history of
both. A critical part of the emerging evolutionary
picture seems to be “horizontal gene transfer” — genetic
change that occurs by the exchange of genetic material
between bacteria. This process allows for sudden
evolutionary leaps that are perhaps not possible through
gradual genetic change and natural selection.

In a paper published in the November 22, 2002 issue of
Science, Blankenship, ASU biochemist Jason Raymond and
colleagues show through a comparative genomic analysis
of five photosynthetic prokaryotic organisms that the
genes that code for the intricate molecular complexes
that perform photosynthesis seem to have originated
through ancient genetic mixing that apparently combined
a variety of independently evolved metabolic processes.

In one of the Astrobiology meeting papers, “Horizontal
Gene Transfer in the Evolution of Nitrogen Fixation,”
Raymond, Blankenship and Rice University’s Janet Siefert
do an analysis of the genomes of a larger group of
bacteria and archaea, comparing in particular similar
genes that code for the protein nitrogenase, a critical
enzyme in nitrogen fixation.

“In the very early earth, there was probably some
available nitrogen in the form of ammonia or something
else, so early life forms didn’t have to fix nitrogen
from the atmosphere. At some point though, things
reached a food crisis — you either find someway to get
the atmosphere’s molecular nitrogen into the cycle or
you die. A minimum input of nitrogen can’t sustain a
big biosphere,” noted Blankenship.

“Nitrogen fixation is one of the most interesting
biological processes because it’s so difficult to do
chemically. Nitrogenase is a very complex enzyme system
that actually breaks molecular nitrogen’s triple bond,”
he said.

The researchers find that similar or “homologous”
nitrogenase genes exist across a broad range of
organisms, and appear to be related to other similar
genes coding for proteins involved in photosynthesis,
as well as to other genes in archaea and bacteria that
do neither photosynthesis nor nitrogen fixation.

“We found a group of homologous genes that doesn’t
correspond to any genes that go with photosynthesis
or any that we know in nitrogen fixation — we found
these in a wide range of organisms,” said Raymond.

The analysis suggests that the related genes that code
for neither nitrogenase nor enzymes in photosynthesis
may be “relics,” coding for metabolic pathways that
are ancestral to both photosynthesis and nitrogen
fixation. Horizontal gene transfer appears to be
responsible for the broad distribution of the original
gene and for its subsequent divergence and
specialization in the metabolic pathways of nitrogen
fixation and photosynthesis.

In the second paper, “The Evolutionary Relationship
between Nitrogen Fixation and Bacteriochlorophyll
Synthesis,” ASU’s Christopher Staples, Blankenship,
and Virginia Polytechnic Institute’s Biswarup
Mukhopadhyay examine the properties of enzymes created
by these similar genes and finds that nitrogenase, the
photosynthesis related enzymes, and other homologous
enzymes all generally belong to a group of enzymes that
break apart molecules and are known as reductase enzymes.

“We’re purifying the proteins that the genes produce and
will be looking at catalytic activity. We will test to
see how activity differs and also to find what has been
conserved and what has been changed in the active sites,”
said Staples. “Changes in the enzymes’ active sites lead
to differentiation in regard to what specific molecules
they affect.”

The less-specialized reductase enzymes appear to be
ancestral to the others and were perhaps originally
important in helping early prokaryotes neutralize
toxic substances in their environment.

“There is a hypothesis that the ancient reductase, in
the presence of a reducing atmosphere, may have been a
hydrogen cyanide reductase,” said Staples.

The team thinks that they have perhaps found a living
model for this in Methanococcus jannaschii, a methane-
producing archaea that performs neither nitrogen
fixation nor photosynthesis but produces a reductase
enzyme that the researchers suspect is used to break
down hydrogen cyanide.

“We’re testing to see if these organisms can grow in
the presence of cyanide and if they can use cyanide
as a nitrogen source,” said Staples. “They don’t
appear to be able to use cyanide exclusively for
nitrogen, but they can grow in concentrations of it
that would be deadly to most organisms.”

While the search to discover the evolutionary history
of the key chemical processes of the biosphere involves
some esoteric genomic and biochemical detective work,
Blankenship, Raymond and Staples point out that
understanding how the chemical processes of photosynthesis
and nitrogen fixation evolved may have some large
practical pay-offs.

“Understanding the origins of nitrogenase, for example,
links to things like the synthesis of fertilizer,” said
Blankenship. “I come from the Midwest where there are
these huge anhydrous ammonia plants that are tremendous
users of energy — a fantastic amount of energy goes
into the making of ammonia. But that’s exactly what
this enzyme complex does: make ammonia out of nitrogen.
It’s a bio solution to this incredibly important and
very expensive process of fertilizer production.

“There’s tremendous appeal in having a bioengineered
version of nitrogen fixation. If we understand this
complex pathway and its origin and evolution, then we
can think more effectively about engineering it into
other places. The benefit to society of being able to
engineer in nitrogen fixation into most crop plants
would be profound,” he said.

Founded in 1998, the NASA Astrobiology Institute
sponsors scientific research in a wide variety of
disciplines aimed at asking and answering large and
fundamental questions about life in the universe.
Astrobiology is an exciting new discipline devoted
to the study of life on Earth and elsewhere in the
universe — its origin, evolution, distribution, and
future. This new area of multidisciplinary study is
bringing together the physical and biological sciences
to address some of the most fundamental questions of
the natural world.

The NASA Astrobiology Institute is composed of over
700 researchers distributed at more that 130 research
institutions across the United States. Its central
offices are located at NASA Ames Research Center, in
the heart of Silicon Valley, California. Additional
information about the NAI can be found at its website:
http://nai.arc.nasa.gov