Restricted by Design

Restricted by Design

Rational Design of Novel Enzyme Highlights Biochemical Design

In 1978, three scientists (Hamilton Smith, Werner Arber, and Daniel Nathans) were awarded the Nobel Prize in Physiology or Medicine for the discovery of restriction enzymes and their applications. These proteins make genetic engineering possible. They have also contributed to the wave of advances that led to the sequencing of the human genome and to the emergence of other biotechnologies.

Restriction enzymes (or endonucleases) are important for another reason. They represent an interesting example of a chicken-and-egg biochemical system and comprise part of the collection of evidence that indicates life must stem from a Creator. Recent work on these proteins highlights this point.

Endonucleases are a family of proteins. This class of biomolecules cleaves DNA. Restriction endonucleases cut both strands of DNA at specific nucleotide sequences, called restriction sites. Specifically, restriction endonucleases protect the cell from foreign DNA, like viruses, by cutting the invading DNA into fragments.

These vital biomolecules occur in conjunction with proteins (called DNA methylases) that attach methyl groups to the same DNA sequences that would normally be cleaved by restriction endonucleases. When these sequences are methylated, restriction endonucleases cannot cut them. Restriction sites of the bacterial DNA are methylated to completely protect the bacterial DNA from being chopped up by its own restriction endonulceases. Foreign DNA, however, is not afforded this same protection.

DNA methylases and restriction endonucleases form a chicken-and-egg pair. Restriction endonucleases would destroy bacterial DNA without DNA methylases. On the other hand, if bacteria did not utilize restriction endonucleases there would be no need for DNA methylases. These two proteins are interdependent and must come into existence simultaneously.

New research by scientists from the Indian Institute of Science (Bangalore, India) helps demonstrate why biochemical systems like restriction endonucleases require the work of an intelligent Agent. These researchers performed experiments to understand the origin of restriction endonucleases from an evolutionary perspective. They also wanted to develop a strategy for engineering novel, nonnatural restriction endonucleases.

Evolutionary biologists think restriction endonucleases evolved from non-specific endonucleases through point mutations in the gene region that codes the DNA binding site on the protein surface. According to this model, once specificity was established recombination and genetic shuffling of the DNA sequences that encode the DNA recognition sites would have generated new restriction endonucleases with different specificities.

To explore this possibility the research team attempted to engineer a highly specific restriction endonuclease from one (R. KpnI) that promiscuously binds to DNA. To accomplish this goal, the scientists employed a rational design strategy to determine which amino acids in the R. KpnI structure to change. These workers had to make use of the detailed understanding of this protein’s structure and functional properties in order to develop the redesign strategy.

They successfully achieved their intended goal by replacing an aspartic acid residue with an isoleucine moiety at amino acid position 163 in the R. KpnI protein chain.

This research illustrates how carefully-thought-through single amino acid substitutions can alter the specificity of restriction endonucleases. This is important work that paves the way to engineer novel, nonnatural restriction enzymes that can expand the arsenal of tools available to molecular biologists and biochemists.

The researchers involved in this study also interpreted their success as support for the evolutionary origin of restriction enzymes with point mutations ushering in the first stage in the molecular evolution of these proteins. At first glance, this interpretation seems warranted.

Still, it’s important to keep in mind that the production of the highly specific restriction endonuclease from the original promiscuous protein required intelligent input from a team of highly trained biochemists who relied on the past work of other highly accomplished scientists. In a sense, this study empirically demonstrates that protein “evolution” requires the work of an intelligent Agent.

It’s also important to note that the researchers didn’t design the companion methylase protein. This protein isn’t necessary for most biotechnology applications. But without the methylase cohort, the reengineered restriction endonuclease would wreck havoc in vivo, destroying DNA that comprises the bacterial genome.

It’s very unlikely that a restriction endonulcease and its partner methylase would simultaneously appear in an evolutionary scenario. These coordinated events would require that changes in the restriction endonuclease would take place at exactly the same time as corresponding changes in the methylase. The only way for coordinated changes like this to happen is under the auspices of an intelligent Agent.

As I point out in my new book The Cell’s Design, human engineers frequently encounter chicken-and-egg problems when designing systems and processes. Everyday experience teaches that chicken-and-egg systems can come to fruition only through intentional planning and implementation. Chicken-and-egg systems, therefore, serve as a potent indicator of intelligent design.

I describe several other examples of chicken-and-egg systems in The Cell’s Design.