Chemical biology probes of mammalian GLUT structure and function.

Biochemical Journal (2018) 475 3511–3534 https://doi.org/10.1042/BCJ20170677 Review Article Chemical biology probes of mammalian GLUT structure and function Geoffrey D. Holman Department of Biology and Biochemistry, University of Bath, Bath BA2 7AY, U.K. Correspondence: Geoffrey D. Holman () The structure and function of glucose transporters of the mammalian GLUT family of proteins has been studied over many decades, and the proteins have fascinated numerous research groups over this time. This interest is related to the importance of the GLUTs as archetypical membrane transport facilitators, as key limiters of the supply of glucose to cell metabolism, as targets of cell insulin and exercise signalling and of regulated membrane traffic, and as potential drug targets to combat cancer and metabolic diseases such as type 2 diabetes and obesity. This review focusses on the use of chemical biology approaches and sugar analogue probes to study these important proteins. Introduction *Geoffrey D. Holman delivered the Biochemical Society’s Sir Philip Randle Lecture in 2017; this review is based on the award lecture given on 7 September 2017. Received: 17 August 2018 Revised: 11 October 2018 Accepted: 11 October 2018 Version of Record published: 20 November 2018 This review is based on the 2017 Randle Lecture delivered at Bath University at a Biochemical Society meeting on ‘Insulin and exercise signalling for glucose homeostasis and metabolic health’. First, I would like to pay tribute to Sir Philip Randle. Sir Philip’s contributions to the field of metabolic control in heath, obesity and type 2 diabetes have been immense. His ideas on the cross-talk between fat and glucose metabolism are still influential today. This review focusses on the first step in glucose metabolism, that is glucose transport, without which cells would not be able to supply metabolic processes with carbohydrate substrates. Sir Philip described the glucose transport step as a site of feedback control of carbohydrate metabolism under conditions in which fat was available as a substrate [1]. This fundamental process has been studied over the many decades using a huge range of techniques and approaches and in this review, I focus on just five of these decades. My career of obsession with, and addiction to, the subject began in 1970 during my PhD work at Southampton University where we worked on sugar analogues that interact with glucose transporters. The single technique that characterises most of our studies is that of chemical biology. We did not describe our analogous studies in the 1970s by this name. This name came into common use much later [2,3]. According to Wikipedia, ‘Chemical biology is a scientific discipline spanning the fields of chemistry, biology, and physics. It involves the application of chemical techniques, tools, and analyses, and often compounds produced through synthetic chemistry, to the study and manipulation of biological systems’. As chemical biology underpinned many of my early studies, and later (from 1976) those of my group at Bath University, I focus this review on these approaches realising, of course, that many researchers in the glucose transport research field have used a wide range of approaches ranging from cell and signalling biology to genetic manipulation and pharmacological interventions. So, what are the glucose transporters that have so fascinated researchers over many decades and what are the key questions concerning their structure and function and regulation? I focus on the mammalian glucose transporters (GLUTs) that catalyse (facilitate) passive movement of glucose down concentration gradients [4]. These gradients are usually from the blood system to the cell interior, but in the liver these gradients can be from the cell to the blood stream. The GLUT family of transport proteins thereby cooperatively function to supply glucose in the direction needed for cell metabolic processes while maintaining a remarkably constant blood glucose level (5 mM after fasting). © 2018 The Author(s). This is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative Commons Attribution License 4.0 (CC BY). 3511 Biochemical Journal (2018) 475 3511–3534 https://doi.org/10.1042/BCJ20170677 The GLUT1 transporter is present in high amounts in human erythrocytes and, because of the relative ease of working with these cells, it has been most studied from the structure related to function perspective [5]. It has a Km value for glucose influx (∼2 mM) which is significantly lower than blood glucose levels [6–8]. The large amounts of the protein (∼5% of the membrane protein) were an important factor in its purification. A peptide sequence was obtained and used to identify a cDNA clone and ultimately the DNA sequence [5]. GLUT1 is still the only endogenous GLUT that has been purified to homogeneity and which can be identified as a Coomassie-staining protein on an SDS–PAGE gel. GLUT1 is present in most human cells and is abundant at the blood–brain barrier [9]. GLUT2 is present in the liver and pancreas and presumably other cells of the endoderm lineage [10]. GLUT3 is mainly present in the brain [11]. A GLUT3 variant (GLUT14) has also been found in the genome as a duplicon of GLUT3 [12], but is essentially uncharacterised and of unknown function and tissue distribution, although there is some disease association [13]. GLUT2 and GLUT3 have Km values for glucose transport that are higher and lower, respectively, than fasting blood glucose reflecting the functions of these proteins in supplying glucose (GLUT2) and rapidly and avidly removing glucose (GLUT3) from the circulation. GLUT4 is present mainly in the insulin-sensitive tissues of adipose, heart and skeletal muscle [14,15]. Its Km for glucose substrate is close to the fasting blood glucose level and this is unchanged by insulin action [16], which instead leads to an increase in glucose transport by increasing GLUT4 translocation to the cell surface of target cells [17]. Genome sequencing has identified 14 mammalian glucose transporter-like proteins which have been divided into three phylogenetically distinct groups [18]. GLUT1–4 and GLUT14 constitute Class 1. The main substrate is glucose with much lower affinity for fructose [19]. GLUTs 5, 7, 9 and 11 are Class 2 transporters. GLUT5 is a fructose transporter with higher affinity for fructose than glucose [20] and is abundant in small intestine, kidney and sperm with lower levels in fat and skeletal muscle [21]. GLUT 7 and 11 transport glucose and fructose with approximately equal affinity [22,23]. GLUT9 has affinity for fructose in the physiological range but also transports urate, and this is probably the physiological substrate [22,24]. The Class 3 transporters include GLUTs 6, 8, 10, 12 and 13, and their tissue distributions, functions and specificities have not been extensively characterised. The preferred substrates for t (...truncated)