Biophysical Reviews’ “meet the editors series”—a profile of Steve Harding’s career in macromolecular hydrodynamics

Abstract

Stephen Harding, Director of the National Centre for Macromolecular Hydrodynamics based in the UK, is invited to introduce himself as a new Executive Editor of Biophysical Reviews.

Dreaming spires and streaming macromolecules

I came into Biophysics like many others via the Physics route. I read Physics at the University of Oxford (a fellow student of class 1973–1976 was Tim Berners-Lee, before he invented the Internet), but what proved most influential and inspirational for my future career was doing a supplementary course in Molecular Biophysics run by Prof Sir David Phillips and Dr. Colin Blake, who previously were a key part of the team that worked out the first crystal structure of an enzyme—lysozyme. Speaking to Colin years afterwards, he said I was still the only Physicist to have ever done their course.

With this, and my other main interest in fluid mechanics, I subsequently took the opportunity to do a PhD with Dr. Arthur Rowe and Dr. Mike Dampier at Leicester developing improved ways of representing the shapes of macromolecules in solution using macromolecular hydrodynamics, extending crude ellipsoid of revolution models with the restriction of two equal axes to the more general tri-axial ellipsoids which dispensed with this restriction—but inevitably involved combinations of different hydrodynamic parameters. Leicester proved a great place to be, particularly with Alex Jeffreys directly upstairs on the 1st floor of our building developing forensic genetics.

Then realising theories needed experimental data to work on, I moved to the University of Bristol to do my first postdoc with JM Creeth the man who—whilst a PhD student at University College Nottingham in the 1940s—discovered hydrogen bonds in DNA using viscometry. I joined him when he was working on the Biophysics of mucins, and there I learnt analytical ultracentrifugation (AUC) and became one of a dwindling band of people still using this fabulous technique invented by Svedberg in the 1920s. Then for my 2nd and final postdoc, I had the honour of working under the guise of another of the AUC pioneers Paley Johnson in the Biochemistry Department at Cambridge, extending my experimental portfolio to light scattering - both “classical” or “static” light scattering and the growing technique of quasi-elastic light scattering which became later known as “dynamic” light scattering. This was before the appearance of the neat and compact instrumentation that adorns almost every Biopharma and crystallisation laboratory but in the days where the whole room was dominated by lasers, collimators, amplifier discriminators and autocorrelators, controlled by BBC Acorn computers rather than PC’s. Sample clarity was so important because of the heavy impact on the scattered signal of small amounts of large impurities present: for this reason, the focus of the work was on macromolecular assemblies such as plant viruses and bacteria/bacterial spore characterisation. Cambridge also gave me the chance to team up with the Fluid Mechanicist John Rallison, and we set up the theory defining the thermodynamic non-ideality behaviour of quasi-rigid macromolecules in solution.

UK National Centre for Macromolecular Hydrodynamics

Then in 1984, the UK government appeared to feel sorry for the large numbers of postdoctoral researchers who were going from one short contract to another and came up with the excellent New Blood Lectureship scheme, to bring new tenured lecturer/researchers into universities and new research. I was lucky enough to get one—helped by the support of Sir Hans Kornberg, head of Biochemistry at Cambridge—which took me to Nottingham, ironically where JM Creeth and his supervisors JM Gulland and DO “Doj” Jordan had done their pioneering DNA work four decades previously. When I joined, all the hydrodynamics had gone, including all the light scattering and analytical ultracentrifuges, so it was a case of start from complete scratch.

With the AUC, it was fairly easy to get set up as it had reached a minimum in popularity, and laboratories were discarding Beckman/Spinco Model E’s or MSE Centriscans all over the UK and Europe, and so helped by a former Beckman engineer, we got set up very quickly (and at virtually no cost) with 2 Model E’s and a Centriscan. A collaboration with Unilever Research provided the funds for dynamic light scattering instrumentation (the Malvern 4700 system). Requests for collaboration came in thick and fast particularly from my former Department in Cambridge, and their new head, Prof Richard Perham, suggested approaching the UK Research Councils for support to set up a National Centre for Macromolecular Hydrodynamics as a fully resourced Facility for Academia and Industry (Fig. 1) — we achieved this in 1987. Our old AUCs were fully modernised with laser light sources and off- and online data capture analysis, and we set up a SEC-MALS instrument from Wyatt-Technology (Santa Barbara, USA) in 1989, a first in Europe for the NCMH. Another first for the NCMH was the acquisition of one of the 1st 4 beta-site models of the new generation Beckman Optima XL-A AUC’s in 1990—this was followed by the acquisition of a Disc Centrifuge from CPS Instruments (Stuart, FL, USA), and all this helped get us established as an International Facility.

Fig. 1.

Steve Harding in the NCMH (May 2022)

Collaboration with German laboratories in the former DDR flourished after reunification in 1989/1990 including presentation of a talk in 2 institutions given in German but with an unfortunate mistake in the title ‘die analytische Ultracentrifuge spinnt wieder’ which translates to ‘the analytical ultracentrifuge goes crazy again’ and not what I thought spins again!

Some of the earlier research highlights are shown in Table 1. Much of the earlier work in developing hydrodynamic and thermodynamic methodology for modelling the shapes and conformations of macromolecules in solution has been featured in Biophysical and Biochemical textbooks such as the Cambridge University Press books Methods in Molecular Biophysics (Serdyuk et al. 2007) and Wilson and Walker's Principles and Techniques of Biochemistry and Molecular Biology (Hofman and Clokie 2018) reflected the re-establishment of hydrodynamic methodology as an important part of macromolecular characterisation portfolio.

Table 1.

Early research highlights

Highlight	Reference
Prediction of solution non-ideality based on protein shape	Rallison and Harding (1985); Harding et al (1998, 1999)
Development of an algorithm for analysing mucins and other complex polydisperse and non-ideal systems using sedimentation equilibrium in the analytical ultracentrifuge	Harding (1985)
Development of triaxial ellipsoids as hydrodynamic models for proteins in solution using ‘hydration independent’ shape function and the ELLIPS suite of algorithms	Harding (1987); Garcia de la Torre and Harding (2013)
Development of SOLPRO algorithm with Prof J. Garcia de la Torre for the bead modelling of complex proteins in solution using ‘hydration independent’ shape functions	Garcia de la Torre et al (1997); Garcia de la Torre and Harding (2013)
Development of a combined SEC-sedimentation equilibrium method for polymer molecular weight distribution analysis	Ball et al (1988, 1990); Harding et al (1991)
Development of off-line automatic data capture of analytical ultracentrifuge patterns	Harding and Rowe (1988)
Development of a simple test for macromolecular heterogeneity in a single sedimentation equilibrium experiment and the MSTAR method for sedimentation equilibrium analysis of polymer molecular weights	Creeth and Harding (1982a,b)
Development and first demonstration of the principle of co-sedimentation in the analytical ultracentrifugation for ligand-macromolecule interactions	Harding (1994); Marsh and Harding (1993); Deacon et al (1998); Harding and Rowe (2010)
First demonstration of a protein which forms trimers – chloramphenicol transacetylase	Harding et al (1987)—see also Morgan and Harding (1997)
First demonstration of a cusp shape conformation for the antibody IgE	Davis et al (1990)—see also Ohlandieck and Harding (2018)
First demonstration of how pegylation can screen antibodies	Lu et al (2008a)
First demonstration of how processing and point mutations can affect the conformation of monoclonal antibodies	Lu et al (2008b)
First application of dynamic light scattering with analytical ultracentrifugation to elucidate the after-process integrity of monoclonal antibodies	Nobbmann et al. (2007)
First demonstration of a weak association in carbohydrate polymers	Patel et al (2007)
First SEC-MALS elucidation of the molecular weight distribution of polysaccharides and mucins	Horton et al (1991); Jumel et al (1996)
First application of dynamic light scattering to bacterial spores and demonstration of resistance to disinfectants	Harding and Johnson (1984); Molina-Garcia et al (1989)
Development of the theory for the concentration dependence of hydrodynamic parameters, tested using TYMV and a more recent extension for the concentration dependence of diffusion	Harding and Johnson (1985a,b); Scott et al (2014)
Development of the technique of flotation equilibrium in the analytical ultracentrifuge	Harding et al (1990)
Development of the linear coil array model for the structure mucins	Harding et al (1983a, b); Harding (1989)
Application of DNA methods to study the extent of Scandinavian ancestry in coastal north-west England	Bowden et al (2008); Harding et al (2010)
Development of ‘Crystallohydrodynamics’: combination of x-ray crystallography with hydrodynamics to elucidate the domain orientation of antibodies	Carrasco et al (2001); Lu et al (2006, 2007)
Resolution of an important discrepancy with a theory relating viscosity determination with molecular shape	Harding et al (1982); Harding (1997)

From carbohydrates that behave like proteins to consolidating the macromolecular structures of Viking ships

More recent research has continued to focus on the development of improved methods for the hydrodynamic characterisation of macromolecules and macromolecular assemblies: improved ways of getting molecular weights—the SEDFIT-MSTAR algorithm (Schuck et al 2014), molecular weight distributions via the “Extended Fujita” and “Multisig” approaches (Harding et al 2011; Gillis et al 2013), conformation (Garcia de la Torre and Harding 2013) and interactions (Harding and Rowe 2010), and concentration dependence of sedimentation and diffusion coefficients (Winzor et al 2021). This has supported work on the nature and stability of glycoconjugate vaccines (Abdelhameed et al 2016a,b; MacCalman et al 2020; Bazhenova et al 2021) and in elucidating the hydrodynamics of microbial resistance mechanisms (Phillips-Jones et al (2017a,b); Phillips-Jones and Harding (2018); Harding (2021)) and the first demonstration of antibiotic-mucin complexation interactions (Dinu et al 2020).

These advances have also led to the discovery of a class of carbohydrate polymer that behave like proteins (Heinze et al 2011; Nikolajski et al 2014; Gross 2014) (Fig. 2) and have allowed us to revisit the 1948 JM Creeth 2-chain model for the structure of DNA (Harding et al 2018; Harding 2019). An offshoot of that work was the use of single nucleotide polymorphisms on Y-DNA linked with surnames to demonstrate high levels of Scandinavian ancestry in coastal North West England (Bowden et al 2008; Harding et al 2010).

Fig. 2.

The carbohydrate polymer AEA1 is one of a class of aminocelluloses which can self-assemble into regular complexes. In this case, AEA1 forms reversible tetramers, which can then form supramolecular structures. These types of self-associative properties are being explored for a number of potential applications, including archaeological wood consolidation in vessels such as the Mary Rose and the Oseberg Viking ship artefacts (Wakefield et al. 2020), alongside other bioinspired polymers. Adapted from Nikolajski et al. (2014).

Our most recent work has focussed on a very novel application of macromolecular hydrodynamics in its use for helping to tailor the generation of bioinspired or ‘green’ biobased polymer consolidants for replacing badly decayed archaeological wood—and in particular the Oseberg Viking ship artefacts—Norway’s national treasure, replacing traditional petro-chemical-based consolidants such as polyethylene glycol. This is a real race against time as some of the artefacts are close to complete disintegration. The new generation consolidants being thoroughly examined include aminocelluloses (Wakefield et al 2020), isoeugenols (McHale et al 2017), lignin extracts (Lu et al 2021), siloxanes (Cutajar et al 2022) and terpene-based polymers (Cutajar et al. 2021).

I conclude by hoping this combination of methodological development in the area of solution biophysics coupled to application to a diverse range of macromolecular systems will be of value to Biophysical Reviews and I would be delighted to help and advise potential authors across a broad area of Biophysics.

Declarations

Conflict of interest

The author declares no competing interests.