Profiling the metabolic proteome of bovine mammary tissue.
2-DE and MALDI mass fingerprinting were used to analyse mammary tissue from lactating Friesian cows. The goal was detection of enzymes in metabolic pathways for synthesis of milk molecules including fatty acids and lactose. Of 418 protein spots analysed by PMF, 328 were matched to database sequences, resulting in 215 unique proteins. We detected 11 out of the 15 enzymes in the direct pathways for conversion of glucose to fatty acids, two of the pentose phosphate pathway enzymes and two of the enzymes for lactose synthesis from glucose. We did not detect enzymes that catalyse the first three reactions of glycolysis. Our results are typical of enzyme detection using 2-DE of mammalian tissues. We therefore advocate caution when relating enzyme abundances measured by 2-DE to metabolic output as not all relevant proteins are detected. 2-D DIGE was used to measure interindividual variation in enzyme abundance from eight animals. We extracted relative protein abundances from 2-D DIGE data and used a logratio transformation that is appropriate for compositional data of the kind represented in many proteomics experiments. Coefficients of variation for abundances of detected enzymes were 3-8%. We recommend use of this transformation for DIGE and other compositional data.
Beddek
Amanda J
AJ
Centre for Biodiscovery and School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand.
Rawson
Pisana
P
Peng
Lifeng
L
Snell
Russell
R
Lehnert
Klaus
K
Ward
Hamish E
HE
Jordan
T William
TW
D000818:Animals; D002417:Cattle; D002952:Citric Acid Cycle; D015180:Electrophoresis, Gel, Two-Dimensional; D005227:Fatty Acids / Q000096:biosynthesis; D005260:Female; D005943:Gluconeogenesis; D005947:Glucose / Q000378:metabolism; D006019:Glycolysis; D007774:Lactation / Q000378:metabolism; D008321:Mammary Glands, Animal / Q000737:chemistry* / Q000201:enzymology; D053858:Metabolic Networks and Pathways; D010427:Pentose Phosphate Pathway; D020543:Proteome / Q000737:chemistry*; D040901:Proteomics / Q000379:methods; D019289:Pyruvic Acid / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D005227:Fatty Acids; D020543:Proteome; D019289:Pyruvic Acid; D005947:Glucose