Biochemists Gain Crystal-Clear Insight into 'Ancient' Enzyme

May 14, 1998

DURHAM, N.C. -- Biochemists from the University of Pennsylvania and Duke University Medical Center have reported analytical studies revealing unexpected new insights into how two very different molecules - a protein and an RNA - work together to form an enzyme that performs one of the fundamental tasks of constructing the protein-making machinery of the cell.

Their findings suggest that the partnership between the two molecules might represent an ancient remnant of an early era in life's evolution when RNA molecules were the enzymatic workhorses of primordial cells, before more versatile proteins evolved to take over the job.

The findings also may offer new targets for antibiotic compounds that could disrupt this key process in bacteria to kill them, the researchers said.

The biochemists reported in the May 1 issue of Science that they had obtained the crystal structure of the protein that is part of the enzyme "ribonuclease P." This enzyme is also known as a "ribozyme," because it is one in which the ribonucleic acid (RNA) functions as a catalyst.

Reporting their work were graduate student Travis Stams and Professor David Christianson of the University of Pennsylvania chemistry department; and Duke Medical Center postdoctoral fellow S. Niranjanakumari and Carol Fierke, associate professor of biochemistry. Their work is supported by the National Institutes of Health.

Ribonuclease P plays a key role in activating a molecule called transfer-RNA (tRNA) after it is first synthesized. Such tRNA molecules are the cellular equivalent of errand boys, latching onto individual subunits of proteins, called amino acids, to carry them to the cell's protein-making machinery, where the amino acids are chemically attached to one another in long stringlike molecules that fold into the cell's working proteins.

Specifically, ribonuclease P is a molecular scissors that helps turn a newly synthesized precursor into a functioning tRNA molecule by snipping off an extraneous segment at one end of the "pre-tRNA" molecule. Similarly, ribonuclease P helps activate another RNA, called ribosomal RNA, that is central to the cell's protein-making machinery.

In their experiments, the University of Pennsylvania researchers produced crystals of the ribonuclease P protein, and obtained the protein's structure through the widely used analytical method of X-ray crystallography. In this technique, an X-ray beam is directed through the protein crystal and is diffracted by the crystal's atoms into a pattern of spots. By analyzing the pattern, chemists can deduce the structure of the protein molecules that make up the crystal.

In analyzing the protein's structure, the biochemists found three regions where the protein could bind RNA. One region had an unusual topology and chemical characteristics that suggested it could be the place where the protein bound the RNA portion of the ribonuclease P enzyme.

However, another RNA-binding region of the protein consisted of a large cleft whose characteristics suggested it grabbed and held the segment of the pre-tRNA that was to be snipped off to produce the functioning tRNA.

These findings were particularly surprising, said the chemists, because most theories held that such proteins played only a structural role, helping fold the RNA into its active form, and not an active role in the catalytic chemical reaction.

"Before we had the structure, we had demonstrated that the protein was very important for binding of the pre-tRNA substrate," Fierke said, "but it wasn't important for binding tRNA. This binding could have been either an indirect effect, like changing the conformation of the RNA, or a direct effect. And in general it's been believed such proteins were only involved in folding the RNA, and not really involved in catalysis.

"But these data suggest that the end of the pre-tRNA snakes through that cleft, with the protein directly contacting the pre-tRNA. This is really a very different mechanism for what these proteins do," said Fierke.

The Duke researchers performed chemical experiments that confirmed this contact, producing altered versions of the protein that would chemically cross-link with the RNA in the cleft, allowing them to unequivocally determine that the protein was holding the RNA.

The Duke researchers also are tinkering with the protein's structure to explore a potential third key RNA binding region on the protein that may help the RNA of the ribonuclease P grab magnesium atoms that it needs to function optimally.

Such findings may offer intriguing insights into how the machinery of living cells first evolved, Fierke said.

"We really didn't expect such a role for the protein," she said. "And, if it turns out to be true, one could speculate that one of the reasons life evolved from an RNA-dependent world to a protein-dependent world is that the RNA required magnesium to do anything. And RNA is not particularly good at forming specific metal binding sites, whereas proteins are particularly good."

The biochemists' work might offer new targets for antibiotics, since ribonuclease P is essential for making all tRNAs, and since the bacterial enzyme is different from the enzyme in higher organisms. Thus, both the Duke and University of Pennsylvania groups propose to design inhibitor compounds that will block some aspect of the critical contacts among the protein, the RNA portion of the ribozyme, and the pre-tRNA.

"There are a great many places one could target to thwart this enzyme," Fierke said.

Duke University

Related Amino Acids Articles from Brightsurf:

Igniting the synthetic transport of amino acids in living cells
Researchers from ICIQ's Ballester group and IRBBarcelona's Palacín group have published a paper in Chem showing how a synthetic carrier calix[4]pyrrole cavitand can transport amino acids across liposome and cell membranes bringing future therapies a step closer.

Microwaves are useful to combine amino acids with hetero-steroids
Aza-steroids are important class of compounds because of their numerous biological activities.

New study finds two amino acids are the Marie Kondo of molecular liquid phase separation
a team of biologists at the Advanced Science Research Center at The Graduate Center, CUNY (CUNY ASRC) have identified unique roles for the amino acids arginine and lysine in contributing to molecule liquid phase properties and their regulation.

Prediction of protein disorder from amino acid sequence
Structural disorder is vital for proteins' function in diverse biological processes.

A natural amino acid could be a novel treatment for polyglutamine diseases
Researchers from Osaka University, National Center of Neurology and Psychiatry, and Niigata University identified the amino acid arginine as a potential disease-modifying drug for polyglutamine diseases, including familial spinocerebellar ataxia and Huntington disease.

Alzheimer's: Can an amino acid help to restore memories?
Scientists at the Laboratoire des Maladies Neurodégénératives (CNRS/CEA/Université Paris-Saclay) and the Neurocentre Magendie (INSERM/Université de Bordeaux) have just shown that a metabolic pathway plays a determining role in Alzheimer's disease's memory problems.

New study indicates amino acid may be useful in treating ALS
A naturally occurring amino acid is gaining attention as a possible treatment for ALS following a new study published in the Journal of Neuropathology & Experimental Neurology.

Breaking up amino acids with radiation
A new experimental and theoretical study published in EPJ D has shown how the ions formed when electrons collide with one amino acid, glutamine, differ according to the energy of the colliding electrons.

To make amino acids, just add electricity
By finding the right combination of abundantly available starting materials and catalyst, Kyushu University researchers were able to synthesize amino acids with high efficiency through a reaction driven by electricity.

Nanopores can identify the amino acids in proteins, the first step to sequencing
While DNA sequencing is a useful tool for determining what's going on in a cell or a person's body, it only tells part of the story.

Read More: Amino Acids News and Amino Acids Current Events is a participant in the Amazon Services LLC Associates Program, an affiliate advertising program designed to provide a means for sites to earn advertising fees by advertising and linking to