New development in the tritium labelling of peptides and proteins using solid catalytic isotopic exchange with spillover-tritium.
The mechanism of the reaction of high temperature solid state catalytic isotope exchange (HSCIE) of hydrogen in peptides with spillover-tritium at 140-180 degrees C was analyzed. This reaction was used for preparing [(3)H]enkephalins such as [(3)H]DALG with specific activity of 138 Ci/mmol and [(3)H]LENK with specific activity of 120 Ci/mmol at 180 degrees C. The analogues of [(3)H]ACTG(4-10) with specific activity of 80 Ci/mmol, [(3)H]zervamicin IIB with specific activity of 70 Ci/mmol and [(3)H]conotoxin G1 with specific activity 35 Ci/mmol were produced. The obtained preparations completely retained their biological activity. [(3)H]Peptide analysis using (3)H NMR spectroscopy on a Varian UNITY-600 spectrometer at 640 MHz was carried out. The reaction ability of amino fragments in HSCIE was shown to depend both of their structures and on the availability and the mobility of the peptide chain. The reaction of HSCIE with the beta-galactosidase from Termoanaerobacter ethanolicus was studied. The selected HSCIE conditions allow to prepare [(3)H] beta-galactosidase with specific activity of 1440 Ci/mmol and completely retained its the enzymatic activity.
Zolotarev
Yu A
YA
Institute of Molecular Genetics, Russian Academy of Sciences, pl. Kurshatova 2, 123182 Moscow, Russia. zolya@img.ras.ru
Dadayan
A K
AK
Bocharov
E V
EV
Borisov
Yu A
YA
Vaskovsky
B V
BV
Dorokhova
E M
EM
Myasoedov
N F
NF
D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000378:metabolism; D002384:Catalysis; D020916:Conotoxins / Q000737:chemistry / Q000378:metabolism; D004743:Enkephalin, Leucine / Q000737:chemistry / Q000378:metabolism; D016308:Enkephalin, Leucine-2-Alanine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D005696:Galactosidases / Q000737:chemistry / Q000378:metabolism; D006859:Hydrogen / Q000737:chemistry; D007553:Isotope Labeling / Q000379:methods*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D054848:Peptaibols; D010455:Peptides / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D011506:Proteins / Q000032:analysis* / Q000378:metabolism; D011869:Radioligand Assay; D051381:Rats; D011957:Receptors, Opioid / Q000378:metabolism; D013696:Temperature; D014316:Tritium / Q000737:chemistry
D020916:Conotoxins; D054848:Peptaibols; D010455:Peptides; D011506:Proteins; D011957:Receptors, Opioid; D014316:Tritium; D004743:Enkephalin, Leucine; D016308:Enkephalin, Leucine-2-Alanine; C029960:conotoxin GI; C077077:zervamicin IIB; D006859:Hydrogen; D005696:Galactosidases; C040996:enkephalin-Leu, Ala(2)-Arg(6)-