scholarly journals Valine oxidation: the synthesis and evaluation of l-[3-3H]valine as a tracer in vivo

1992 ◽  
Vol 68 (1) ◽  
pp. 139-151 ◽  
Author(s):  
P. R. Beckett ◽  
A. Cadenhead ◽  
M. F. Fuller
Keyword(s):  
Nuclear Magnetic Resonance ◽  
Amino Acid ◽  
Magnetic Resonance ◽  
Ion Exchange ◽  
Oxidation Rate ◽  
Tritium Atom ◽  
Significant Difference ◽  
Amino Acid Requirements ◽  
Acetyl Group

The suitability of l-[3-3H]valine for measuring valine oxidation was studied by comparing its oxidation rate with that of l-[1-14C]valine in rats and pigs. l-[3-3H]valine was synthesized by removal of the tritium on carbon-2 of l-[2,3-3H]valine by acetylation. The acetyl group was removed enzymatically using pig renal acylase 1 (EC 3.5.1.14) and the product was purified by ion-exchange and paper chromatography. For the first rat experiment l-[3-3H]valine was synthesized in our laboratory; for the subsequent experiments it was produced by Amersham International plc. In the first experiment in rats the two tracers were given by injection and 14CO2 was collected for 2 h. The oxidation of tritiated valine was significantly higher than that of l-[1-14C]valine. In a second experiment there was no difference. This was probably due to the higher purity of the labelled valine which, for the second experiment, was shown by nuclear magnetic resonance to contain only one tritium atom. In a study with pigs in which the two tracers were given by continuous infusion there was no significant difference between them in flux or oxidation. The results of this experiment were used to evaluate a model to estimate amino acid requirements. With pigs given a methionine-limiting diet a reduction in methionine intake, by reducing protein accretion, increased valine oxidation by the same proportion.

In vivo incorporation of isotopically labelled amino acids into vertebrate proteins: L-[Me-3H]methionine incorporation into chicken egg white lysozyme

1977 ◽  
Vol 55 (4) ◽  
pp. 439-444 ◽  
Author(s):  
Kathleen D. Chance ◽  
Donald G. Clark
Keyword(s):  
Amino Acids ◽  
Nuclear Magnetic Resonance ◽  
Amino Acid ◽  
Magnetic Resonance ◽  
Free Amino Acid ◽  
Free Amino ◽  
Egg White ◽  
Magnetic Resonance Studies ◽  
Egg White Lysozyme

The in vivo incorporation of L-[Me-3H]methionine into egg white lysozyme of the laying hen has been examined. Using a versatile synthetic chicken diet which consisted of 75% free amino acid ration and 25% normal laying ration, a 5–8% incoiporation of the [3H]methionine into lysozyme was demonstrated. The utility of this vertebrate in vivo incorporation technique is discussed in terms of its application to the incorporation of 13C-enriched amino acids into vertebrate proteins as a prelude to macromolecular 13C nuclear magnetic resonance studies.

scholarly journals Identification of the Yeast R-SNARE Nyv1p as a Novel Longin Domain-containing Protein

2006 ◽  
Vol 17 (10) ◽  
pp. 4282-4299 ◽  
Author(s):  
Wenyu Wen ◽  
Lu Chen ◽  
Hao Wu ◽  
Xin Sun ◽  
Mingjie Zhang ◽  
...  
Keyword(s):  
Nuclear Magnetic Resonance ◽  
Amino Acid ◽  
Magnetic Resonance ◽  
Adaptor Protein ◽  
Resonance Spectroscopy ◽  
Amino Acid Substitutions ◽  
Sorting Signals ◽  
Distinct Role

Using nuclear magnetic resonance spectroscopy, we establish that the N-terminal domain of the yeast vacuolar R-SNARE Nyv1p adopts a longin-like fold similar to those of Sec22b and Ykt6p. Nyv1p is sorted to the limiting membrane of the vacuole via the adaptor protein (AP)3 adaptin pathway, and we show that its longin domain is sufficient to direct transport to this location. In contrast, we found that the longin domains of Sec22p and Ykt6p were not sufficient to direct their localization. A YXXΦ-like adaptin-dependent sorting signal (Y31GTI34) unique to the longin domain of Nyv1p mediates interactions with the AP3 complex in vivo and in vitro. We show that amino acid substitutions to Y31GTI34 (Y31Q;I34Q) resulted in mislocalization of Nyv1p as well as reduced binding of the mutant protein to the AP3 complex. Although the sorting of Nyv1p to the limiting membrane of the vacuole is dependent upon the Y31GTI34 motif, and Y31 in particular, our findings with structure-based amino acid substitutions in the mu chain (Apm3p) of yeast AP3 suggest a mechanistically distinct role for this subunit in the recognition of YXXΦ-like sorting signals.

Biological NMR Spectroscopy

1997 ◽  
Keyword(s):  
Nuclear Magnetic Resonance ◽  
Nmr Spectroscopy ◽  
Magnetic Resonance ◽  
Magnetic Resonance Spectroscopy ◽  
State Of The Art ◽  
Critical Assessment ◽  
Resonance Spectroscopy ◽  
History Of ◽  
Structure Of Proteins

This book presents a critical assessment of progress on the use of nuclear magnetic resonance spectroscopy to determine the structure of proteins, including brief reviews of the history of the field along with coverage of current clinical and in vivo applications. The book, in honor of Oleg Jardetsky, one of the pioneers of the field, is edited by two of the most highly respected investigators using NMR, and features contributions by most of the leading workers in the field. It will be valued as a landmark publication that presents the state-of-the-art perspectives regarding one of today's most important technologies.

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