X-rays Of Cell's Ribosome Could Lead To Better Antibiotics

[Guest guest]

Source: University of California - Berkeley

Date: 2005-11-06

URL: http://www.sciencedaily.com/releases/2005/11/051106181101.htm

New And Sharper X-rays Of Cell's Ribosome Could Lead To Better

Antibiotics

A new, sharper picture of the nano-machine that translates our

genetic program into proteins promises to help researchers explain

how some types of antibiotics work and could lead to the design of

better ones.

The high-resolution snapshots of the bacterial ribosome were

captured by scientists at the University of California, Berkeley,

and Lawrence Berkeley National Laboratory (LBNL) with the lab's

Advanced Light Source, which generates intense beams of X-rays that

can reveal unprecedented structural detail of such large and complex

molecules.

The new, high-resolution data on the intact ribosome allows

researchers to build more detailed and more realistic models of the

ribosome that until now were impossible with the " fuzzy pictures "

available.

While sharp images of the two main pieces of the ribosome have

already provided great insight into how specific antibiotics work,

many antibiotics, such as the aminoglycosides, only interfere with

the entire, fully assembled molecular machine.

" Many antibiotics target only the intact machine, disrupting

messenger RNA decoding or movement, " said lead author Cate,

assistant professor of chemistry and of molecular and cell biology

at UC Berkeley and a staff scientist in the Physical Biosciences

Division at LBNL. " We are now in a position to look at some of these

drugs and discover things that haven't been known before. "

Cate, a member of the California Institute for Quantitative

Biomedical Research (QB3) at UC Berkeley, and his colleagues report

the detailed structure of the ribosome from Escherichia coli, the

common intestinal bacteria, in the Nov. 4 issue of Science.

The ribosome, about 21 to 25 nanometers across, is the original

nanomachine, taking genetic information relayed by messenger RNA,

decoding it and spitting out proteins. Ribosomes are dispersed in

the hundreds of thousands throughout the cell, and in some highly

active cells, ribosomes are responsible for producing millions of

proteins per minute.

Ribosomes are found in all organisms, ranging from bacteria to

humans, and probably arose nearly 2 billion years ago. They have

changed so little through evolution that a bacterial ribosome can

often translate human genes into protein. Some people suspect that

ribosomes, which at their core consist of ribonucleic acid (RNA), a

sister of the DNA that comprises our genes, arose when RNA, not DNA,

carried our genetic dowry.

Because of its importance to life, and the fact that important drugs

target the ribosome, it has received lots of attention. Only four

years ago, Cate was part of a team that published a picture of the

ribosome with a resolution of 5.5 Angstroms, where an Angstrom,

about the size of a hydrogen atom, is one-tenth of a nanometer. The

new images have a resolution of 3.5 Angstroms, allowing Cate and his

colleagues to see the individual nucleotides in the RNA strands of

the ribosome and the amino-acid backbones of the proteins that

surround the RNA core.

Both the old and new images were obtained through X-ray

crystallography using Advanced Light Source beamlines, which provide

extremely bright X-ray sources. Having the light source in his

backyard, Cate said, has made it easier to get the best

crystallographic picture with the sharpest three-dimensional detail.

He and his laboratory colleagues grow crystals of ribosomes, check

their quality in the light source, then tweak the crystals and try

again.

" We've burned through thousands of crystals in the last five years, "

he said.

The researchers obtained two high-resolution snapshots of the intact

E. coli ribosome and compared them with a wide range of

conformations of other ribosomes. These other data came from lower-

resolution X-ray crystallographyic images of Thermus thermophilus

and E. coli ribosomes, plus electron microscopy of E. coli, yeast

and mammalian ribosomes. Together, they yielded what Cate

calls " global snapshots " and allowed him and his colleagues to

deduce how individual parts of the ribosome function during the

translocation process.

What the new structure shows so far is how the two large pieces of

the ribosome bend, ratchet and rotate as the ribosome goes through

the repetitive process of protein manufacturing.

The " small " subunit of the ribosome first recognizes and latches

onto the messenger RNA (mRNA), which contains a copy of part of the

chromosomal DNA. Once the small subunit finds the start position,

the " large " subunit moves in and latches on, clamping the mRNA

between them. The combined machine slides along the mRNA, reading

each three-letter codon, matching this code to the appropriate amino

acid, and then adding that amino acid - one of 20 possible building

blocks - to the lengthening protein chain.

As this translation takes place, transfer RNA (tRNA) constantly

brings in amino acid building blocks, while energy-supplying

molecules in the form of GTP (guanosine triphosphate) cycle through.

They found that after the bond - called a peptide bond - forms

between the growing chain and the newly added amino acid, the small

subunit ratchets with respect to the large subunit. Then the head of

the small subunit swivels in preparation for shifting the mRNA

forward by one codon. At the same time, a groove opens that allows

the mRNA to actually move and the tRNA, depleted of its amino acid,

to float away.

Then, the small subunit reverses its motions, resets, and is ready

to add the next amino acid. This picture of translocation -

ratcheting, swiveling, opening the groove, then reversing these

three steps - is repeated 10 to 20 times each second in bacteria.

Based on the researchers' analysis of the new data, Cate said that

it appears, also, that the helical RNA in the ribosome acts as a

spring to withstand the stress of these reversible swivels. Also,

the ribosome harbors an astounding number of positive magnesium

ions - hundreds in all - that apparently neutralize the highly

negative charge of the RNA. Without these magnesium ions, Cate said,

the repulsion of the RNA's negative charge would blow the ribosome

apart. Some of the magnesium ions form a salty liquid at the

interface between the large and small subunits of the ribosome,

perhaps lubricating the machine.

These and other hypotheses need further exploration, he said.

" All the interactions we see have been seen before at lower

resolution, but it was not clear how to interpret them, " he

said. " It took these high-resolution studies to coalesce our ideas. "

###

The study's coauthors include Barbara S. Schuwirth in UC Berkeley's

Department of Chemistry and the Free University of Berlin's

Institute of Chemistry-Crystallography; A. Borovinskaya, Antón

Vila-Sanjurjo and M. Holton of LBNL; and W. Hau and Wen

Zhang of UC Berkeley's Department of Molecular and Cell Biology.

The work was supported by the National Institute of General Medical

Sciences and the National Cancer Institute of the National

Institutes of Health and by the U.S. Department of Energy.