ACPFG Blog

Making membrane proteins: curdlan synthase

December 8, 2012

Filed under: Research — Tags: , — acpfg @ 8:43 am

Curdlan is a polysaccharide which is used in Chinese medicine to stimulate the immune system to prevent skin infections. Researchers at the Australian Centre for Plant Functional Genomics have managed to make curdlan synthase, the enzyme that synthesises curdlan, in the laboratory.

The techniques they used to produce the enzyme should be very useful to finding out about some of nature’s most secretive proteins – membrane proteins.

Scientists have solved only around 350 membrane protein structures compared to more than 84 000 structures of soluble proteins. And membrane proteins are some of the most important proteins in biology – for example about half of all medications achieve their effects through membrane protein receptors. In humans, membrane proteins detect signals from hormones like adrenaline, and in plants they produce cellulose and other polysaccharides that build cell walls or protect plant cells.

Diagram showing membrane proteins of different types embedded in the cell membrane

There are several types of membrane proteins. They are hard to produce in the laboratory because they’re not stable unless they’re bound to lipids. Maria Hrmova’s group uses surfactants during protein synthesis to stabilise the proteins.
Image: modified from a public image.

Despite their importance, membrane proteins are notoriously difficult to produce in the laboratory. Most protein structural biologists produce proteins by getting bacteria like E. coli to do the job. They give the bacteria the relevant gene, and use a series of chemical tricks to get them to make protein. But bacteria aren’t quite like plants or animals, and that means they can’t always make plant and animal proteins successfully.

When Maria Hrmova and her team at ACPFG used E. coli bacteria to make the membrane protein curdlan synthase, the bacteria didn’t do the job properly. The bacteria produced parts of the protein, but never its full length. Bacteria often have trouble making membrane proteins because they need to be produced in a special way so they’re protected by membranes as they’re made.

So Maria and her team turned away from bacteria, and used an extract from plant cells combined with lipids and surfactants to produce the protein. And the method worked. The curdlan synthase they made was full length and it was correctly synthesised in artificial membranes made of lipid bilayers. They could also reconstitute the protein in lipid nano-discs. Most importantly, the protein was happy to produce curdlan.

Next Maria and her team will be trying to crystallise the protein so they can find out its three-dimensional molecular structure and get the details of how the enzyme joins individual sugars together to make curdlan. The scientists hope this will contribute to research into curdlan’s medical uses. It will also add to our knowledge of how plants create their cell walls.

The research was published in BBA-Biomembranes.

The scientists who worked on the project were Agalya Periasamy, Nadim Shadiac, Amritha Amalraj, Soňa Garajová, Yagnesh Nagarajan, Shane Waters, Haydyn D.T. Mertens and Maria Hrmova.

This article was written by Arwen Cross and the image was prepared by Maria Hrmova.

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