NMR and Microorganisms

Abstract

This article is an introduction to the use of NMR for the investigation of microbial physiology and metabolism. NMR parameters which determine the sensitivity and resolving power of the method are reviewed. A broad survey of current applications follows. Qualitative uses are described first; they include compound identification and localisation. Quantitative aspects, such as pH, concentration and flux measurements are then examined, as well as the corresponding experimental constraints. The review ends with suggestions of possible future developments in instrument capabilities aimed at improving sensitivity: higher fields, spectroscopic and imaging microprobes. IntroductionThe first report on the application of NMR to microorganisms is that of Eakin et al. on the yeast, Candida utilis (1). During the past 27 years, many developments have taken place, which make in vivo NMR a very useful tool in the study of microbial physiology and metabolism. It is the purpose of this introductory review to provide the reader with an orientation to the field. It is comprised of four parts. We first list a few characteristics of the NMR technique which are deemed important for metabolic investigation. We then describe several applications of NMR, first as a qualitative tool then as a quantitative instrument. We have taken the liberty to mention advances which have taken place in related fields, such as biofluid analysis and NMR studies of mammalian cells or whole organs, whenever they were perceived as useful and readily adaptable to the case of micro-organisms. We end by mentioning various probable or desirable future developments. Some Aspects of NMR of Importance in MicrobiologyWe assume that the reader has some knowledge of NMR. We will be mainly concerned with small molecules, observed either in homogenous solutions (supernatant or growth medium, cell extracts) or inside living cells. SensitivityThe sensitivity of the NMR experiment should probably be the foremost concern of prospective users. It depends on the following factors. 1) Nature of the nucleus: Each magnetic nucleus is characterised by a spin quantum number I > 0 and a gyromagnetic ratio γ. Several elements which play an important role in biochemistry have "low γ" nuclei (Mg, Ca) and present special problems. A further point is that nuclei with I $1 (such as 14 N) have a quadrupole moment and display very broad lines when bound in a molecule. Exceptions include alkali, alkaline earth, and chlorine ions, provided they are not bound to a carrier macromolecule. 2) Operating magnetic field: The sensitivity depends roughly on the square of the static field. As the price of a spectrometer is also a steep function of the magnetic field, there is an obvious economic limitation here. It is compounded with the requirement of a wide sample access, because magnet prices also increase with sample bore. 3) Active volume: Provided the sample has a correct shape and is relatively homogenous, sensitivity is roughly proportional to the volume inside the radio-frequency coi...

Metabolic Plasticity of Candida albicans in Response to Different Environmental Conditions

“…Recognizing that metabolic profiling provides a direct “functional readout of the physiological state of an organism” [ 26 ], an increasing number of metabolomics-based studies have contributed to further understanding the molecular mechanisms that govern the microorganisms’ life [ 27 , 28 , 29 ]. Yeasts represent the best eukaryotic model, and comparative analyses of their metabolic profiles under different experimental conditions, in the presence of drugs, or when using genetically manipulated strains have provided precious hints at the molecular level [ 30 , 31 , 32 , 33 , 34 , 35 , 36 , 37 , 38 , 39 , 40 ].…”