Degradation of structurally characterized proteins injected into HeLa cells. Basic measurements

Journal Article · Sun Dec 25 04:00:00 EST 1988 · J. Biol. Chem.; (United States)

OSTI ID:6786454

Rogers, S W; Rechsteiner, M

Thirty-five proteins of known x-ray structure were labeled by chloramine-T radioiodination or by reaction with 125I-Bolton-Hunter reagent and introduced into HeLa cells using red cell-mediated microinjection. Degradation rates of the injected proteins were then determined over the next 50 h by measuring the release of soluble isotope to the culture medium. Control experiments demonstrated that the measured rates were not compromised by proteolysis within RBCs, the presence of unfused RBCs, or degradation of protein released from RBCs to the medium. Degradation of some injected proteins was faster during the first 12 h after fusion than at later times, apparently a response of HeLa cells to trypsinization. However, all proteins exhibited first-order degradation rates between 24 and 48 h post injection. Except for seven proteins, stabilities measured during this interval were unaffected by the labeling procedure. Reductive methylation was used to choose among the seven discordant values, and half-lives for the 35 proteins ranged from 16 h for lysozyme to 214 h for yeast alcohol dehydrogenase. Since half-lives for six of the injected proteins closely match values obtained by in vivo measurements, we consider our estimates of the metabolic stabilities of the injected proteins to be generally accurate. Therefore, the half-lives obtained by microinjection should prove useful in the search for relationships between protein structure and intracellular stability.

• Research Organization: Univ. of Utah, Salt Lake City (USA)
• OSTI ID: 6786454
• Journal Information: J. Biol. Chem.; (United States), Journal Name: J. Biol. Chem.; (United States) Vol. 263:36; ISSN JBCHA
• Country of Publication: United States
• Language: English

Related Subjects

550501* -- Metabolism-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AUTORADIOGRAPHY
BETA DECAY RADIOISOTOPES
BIOLOGICAL HALF-LIFE
BIOLOGICAL MATERIALS
BLOOD
BLOOD CELLS
BODY FLUIDS
COHERENT SCATTERING
DAYS LIVING RADIOISOTOPES
DIFFRACTION
ELECTRON CAPTURE RADIOISOTOPES
ERYTHROCYTES
HELA CELLS
INTERMEDIATE MASS NUCLEI
IODINE 125
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPE DILUTION
ISOTOPES
MATERIALS
METABOLISM
MOLECULAR STRUCTURE
NUCLEI
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
PROTEINS
RADIOISOTOPES
SCATTERING
STABILITY
TRACER TECHNIQUES
X-RAY DIFFRACTION