Understanding the molecular with biochemistry and biophysics

By Emma Pettengale, Commissioning Editor, Portland Press

Why the molecular?

Molecular biosciences explore the structure and function of biomolecules within your cells and the processes that allow cells to live, reproduce and communicate with each other. Biomolecules are the building blocks for all life on Earth, from the simplest viruses, through bacteria to complex eukaryotic organisms and underpin the processes of transcription, translation, replication and cell function.

What is biophysics?

According to the Biophysical Society, “biophysics is a bridge between biology and physics”, and offers the perspective that biological processes can be understood from the interactions between and within the constituent molecules. This field of study investigates the molecular biosciences using methods based on principles of physics, including light, X-ray or other electromagnetic spectrum scattering; fluorescence methods, including FRET; changes in voltage or currents; and microscopy and imaging techniques.

What is biochemistry?

Similarly, biochemistry seeks to understand how biological molecules give rise to the processes that occur within living cells, which in turn relates greatly to the study and understanding of tissues, organs, and whole organisms. Biochemists explore the chemical process within living organisms using chemical methods such as changes in target concentrations and chemical reactions between molecules; the structure and function of macromolecules such as proteins and DNA; inorganic ion chemistry; and the processes that allow our cells to use chemical energy to function.

What are the questions they both seek to answer?

The line between the biophysical and the biochemical have become blurred, with some research labs drawing from both fields to explore the molecular biosciences.

Questions scientists in both fields seek to answer include:

  • What structures do RNA and DNA form? What role does the structure of nucleic acid play in its function? Carika Weldon, Ian Eperon and Cyril Dominguez explores whether long RNA molecules can from G-quadruplex structures in this recent open access review.
  • How is DNA replicated faithfully and how do cells repair errors in the genetic code?
  • How are biomolecules made in the cell?
  • How do nascent polypeptides fold? Check out this open access review from the Biochemical Journal if you’re interested in protein folding fundamentals and mechanisms from ultrafast approaches

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Protein folding inside the cell (Munoz & Cerminara, Biochemical Journal 2016 DOI: 10.1042/BCJ20160107)

  • How are membranes structured? How do they assemble and do we fully understand their function? This open access paper explores the different methods used to study the oligomeric structure of G protein-coupled receptors – both biochemical and biophysical
  • How do we measure cellular metabolic processes? Find out more about this in a review from Nealon and Howard, which explores using NMR spectroscopy.
  • The dynamic structure of the cell is maintained by the cytoskeleton – how does the cell maintain and regulate this complex system during development, stasis, cell division and motility? How is this process disrupted in cancer metastases?
  • How do plants photosynthesise and can we apply their methods to create better fuels and technology to power our world?
  • How do viruses and bacteria infect our cells and how does the immune system respond at the cellular level?
  • Some diseases are caused by errors at the molecular level, can we identify them and develop therapies to correct them and thus treat the disease?

At the 61st Annual Biophysical Society Meeting held last month in New Orleans, USA, sessions and posters presented research across a range of topics seeking to answer some of these fundamental questions, including focuses on protein structure, membrane fusion, membrane structure, ion channels and the cytoskeleton.

As we celebrate Biophysics Week 2017, we would like to highlight some research from the Biochemical Society journals exploring these shared themes which might be of interest.

Protein structure relates directly to function, both biophysics and biochemistry aims to elucidate those structures and provide insight into important functions.

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Interaction of PS1 with hArc (CC-BY Myrum et al. Biochemical Journal 2015. DOI: 10.1042/BJ20141446)

In their recent research article, Craig Myrum et al. expressed and purified recombinant human Arc (hArc) and performed the first biochemical and biophysical analysis of the protein’s structure and stability. They began with the in silico prediction of secondary structure from the amino acid sequence and used biochemical assays, including limited proteolysis, to show that hArc is a protein with two domains, before seeking to confirm the functionality of recombinant hArc by its ability to associate with PS1, by comparing the CD spectra for hArc and PS1, both separate and together (see Figure 2).

 

 

 

They conclude that hArc is a flexible multi-domain protein that exists in monomeric and oligomeric forms, compatible with a diverse, hub-like role in plasticity-related processes.  If you would like to find out more about Arc, you can also check out this associated commentary from Cameron Day and Jason Shepherd at the University of Utah.

Ezrin is a protein with dormant and active forms, where the latter links the actin cytoskeleton to membranes, and which forms both stable monomers and dimers. Phang et al used SAXS (small-angle X-ray scattering) to determine the solution structures of these species and found that the monomer shows a globular domain with a protruding helical coiled-coil while the dimer shows an elongated dumb-bell structure, twice as long as the monomer.

 

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Structural plasticity in the core of subdomain F3 in the ezrin FERM domain structure (Biochemical Journal 2016 DOI: 10.1042/BCJ20160541)

A key question both biochemists and biophysicists try to answer is how do proteins fold? In this open access review, Horne and Radford discuss old, new and emerging techniques used to examine the process of β-barrel outer membrane protein folding and biogenesis and describe some of the insights and new questions these techniques have revealed (Figure 4).

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‘Classic’ methods of interrogating protein folding Different techniques provide independent, and complementary, information about the kinetics, thermodynamics and mechanism of folding. (CC-BY 4.0 Horne & Radford, Biochemical Society Transactions DOI: 10.1042/BST20160020)

In addition to regulating proteins via biogenesis, cells also carefully control their destruction to ensure quality control and appropriate activity. One of the key aspects of this regulation, known as proteostasis, is the disposal of misfolded proteins by, organelles that are responsible for the breakdown of proteins and other cellular components. In amyloid diseases, such as Parkinson’s disease, there is a failure in proteostasis in which proteins misfold into amyloid aggregates (see Figure 5). You can find out more about proteostasis in this themed issue with guest editor Patricija van Oosten-Hawle.

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Cellular pathways of protein degradation remove misfolded proteins (CC-BY 4.0 Essays in Biochemistry 2016 DOI: 10.1042/EBC20160005)

As well as performing structural and metabolic processes within the cell, proteins are also found in membranes.

SNAREs are a highly conserved set of membrane-associated proteins that mediate intracellular membrane fusion. In this open access review, Lou and Shin describe some recent progress in understanding the pathway of SNARE zippering.

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A structural model of the half-zippered intermediate: (A) Trans-SNAREpin representing the fusogenic pre-fusion intermediate trapped between two nanodisc membrane patches; (B) The low FRET population represents the half-zippered species whereas the high FRET population reflects the fully zippered species (CC-BY 4.0 Bioscience Reports 2016, 36 (3) e00327; DOI: 10.1042/BSR20160004)

Membrane structure and function is a key area of interest for all molecular bioscientists, cells exploit a wide range of physical and biochemical mechanisms to modulate lipid and protein localization spatially and temporally on membranes.

In this review, Byrum & Rodgers discuss how techniques that resolve nanoscale clustering of membrane proteins, such as FRET and electron microscopy, show that disrupting the cytoskeleton can be as effective as reducing plasma membrane cholesterol towards de-clustering cholesterol-dependent membrane-associated proteins. This figure (Figure 7) shows the rationale for the FRET-based approach for measuring lipid ordering in nanodomains.

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Detection of nanoscopic membrane heterogeneities by FRET (Essays in Biochemistry Feb 15, 2015, 57 177-187; DOI: 10.1042/bse0570177)

EP - pic11Ion channels and protein (nano)pores play a key role in membrane biology, allowing rapid movement of ions and/or water molecules across cell membranes, while gating is the switching of a channel between an ‘open’ state which enables permeation and a ‘closed’ sate which prevents permeation. One possible mechanism for closure is ‘hydrophobic gating’, this review explores recent simulation studies of hydrophobic gating in simple model systems.

Schematic diagram (Right) illustrating hydrophobic gating, a ‘vapour lock’ (Biochemical Society Transactions 2015, 43 (2) 146-150; DOI: 10.1042/BST20140256)

When exploring biological processes such as protein synthesis, a biochemist may be interested in reducing the process into a sequence of binding events and chemical reactions, whereas a biophysicist may be interested in the rate constants of these events. So biochemistry identifies the biomolecules involved, and biophysics can then explain the biochemical observations based on the structures and the interactions of the constituent molecules. Hence, exploring the molecular mechanisms behind processes such as replication, transcription, translation and cell function, biophysicists and biochemists approach the same questions from different angles, neither able to provide the full picture but both are needed to discover how the process actually works. 

Many biochemists are also biophysicists, “In actual day to day research you’re less concerned about if what you’re doing is biochemistry or biophysics and more concerned about if you can answer your question.”

If you’d like to find out more about any of the research mentioned above, or are interested in learning more about molecular bioscience and the links between biochemistry and biophysics check out the combined collection of interesting papers from Portland Press!

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