Prediction of growth with kinetics of cell-free translation systems

1985 ◽  
Vol 70 (6) ◽  
pp. 667-670 ◽  
Author(s):  
B. Loehrke
Keyword(s):  
Free Translation ◽  
Translation Systems ◽  
Kinetics Of

scholarly journals Context effects and inefficient initiation at non-AUG codons in eucaryotic cell-free translation systems.

1989 ◽  
Vol 9 (11) ◽  
pp. 5073-5080 ◽  
Author(s):  
M Kozak
Keyword(s):  
Context Effects ◽  
Consensus Sequence ◽  
Negative Regulator ◽  
Late Event ◽  
Free Translation ◽  
Initiator Codon ◽  
Translation Systems ◽  
Short Versions

The context requirements for recognition of an initiator codon were evaluated in vitro by monitoring the relative use of two AUG codons that were strategically positioned to produce long (pre-chloramphenicol acetyl transferase [CAT]) and short versions of CAT protein. The yield of pre-CAT initiated from the 5'-proximal AUG codon increased, and synthesis of CAT from the second AUG codon decreased, as sequences flanking the first AUG codon increasingly resembled the eucaryotic consensus sequence. Thus, under prescribed conditions, the fidelity of initiation in extracts from animal as well as plant cells closely mimics what has been observed in vivo. Unexpectedly, recognition of an AUG codon in a suboptimal context was higher when the adjacent downstream sequence was capable of assuming a hairpin structure than when the downstream region was unstructured. This finding adds a new, positive dimension to regulation by mRNA secondary structure, which has been recognized previously as a negative regulator of initiation. Translation of pre-CAT from an AUG codon in a weak context was not preferentially inhibited under conditions of mRNA competition. That result is consistent with the scanning model, which predicts that recognition of the AUG codon is a late event that occurs after the competition-sensitive binding of a 40S ribosome-factor complex to the 5' end of mRNA. Initiation at non-AUG codons was evaluated in vitro and in vivo by introducing appropriate mutations in the CAT and preproinsulin genes. GUG was the most efficient of the six alternative initiator codons tested, but GUG in the optimal context for initiation functioned only 3 to 5% as efficiently as AUG. Initiation at non-AUG codons was artifactually enhanced in vitro at supraoptimal concentrations of magnesium.

Acetyl Phosphate as an Energy Source for Bacterial Cell-Free Translation Systems

1995 ◽  
Vol 226 (1) ◽  
pp. 184-186 ◽  
Author(s):  
L.A. Ryabova ◽  
L.M. Vinokurov ◽  
E.A. Shekhovtsova ◽  
Y.B. Alakhov ◽  
A.S. Spirin
Keyword(s):  
Energy Source ◽  
Bacterial Cell ◽  
Acetyl Phosphate ◽  
Free Translation ◽  
Translation Systems

scholarly journals Cell-free translation systems prepared from starfish oocytes faithfully reflect in vivo activity; mRNA and initiation factors stimulate supernatants from immature oocytes.

1990 ◽  
Vol 1 (13) ◽  
pp. 1057-1067 ◽  
Author(s):  
Z Xu ◽  
M B Hille
Keyword(s):  
Translational Control ◽  
Translational Regulation ◽  
High Capacity ◽  
Meiotic Maturation ◽  
Immature Oocytes ◽  
Initiation Factors ◽  
Initiation Of Translation ◽  
Free Translation ◽  
Translation Systems

Meiotic maturation stimulates a change in the translation of stored mRNAs: mRNAs encoding proteins needed for growth of oocytes are translated before meiotic maturation, whereas those encoding proteins required for cleavage are translated after meiotic maturation. Studies of translational regulation during meiotic maturation have been limited by the lack of translationally active cell-free supernatants. Starfish oocytes are ideal for preparing cell-free translation systems because experimental application of the hormone 1-methyladenine induces their maturation, synchronizing meiosis. We have prepared such systems from both immature and mature oocytes of starfish. Changes in protein synthesis rates and the specificity of proteins synthesized in these cell-free translation supernatants mimic those seen in vivo. Supernatants both from immature and mature oocytes have a high capacity to initiate new translation because 90% of the proteins made are newly initiated from mRNAs. Cell-free supernatants from mature oocytes have a much higher rate of initiation of translation than those from immature oocytes and use the 43S preinitiation complexes more efficiently in initiation of translation. Similarly, we have shown that mRNAs and initiation factors are rate limiting in cell-free translation systems prepared from immature oocytes. In addition, cell-free translation systems prepared from immature oocytes are only slightly, if at all, inhibitory to cell-free translation systems from mature oocytes. Thus, soluble inhibitors, if they exist, are rapidly converted by cell-free supernatants from mature oocytes. The similarities between translation in our starfish cell-free translation systems and in intact oocytes suggests that the cell-free translation systems will be useful tools for further studies of maturation events and translational control during meiosis.

Yeast Cell-Free Translation Systems

2002 ◽  
pp. 67-78 ◽  
Author(s):  
Michael Altmann ◽  
Hans Trachsel
Keyword(s):  
Yeast Cell ◽  
Free Translation ◽  
Translation Systems

Fluorescence labeling of the C-terminus of proteins with a puromycin analogue in cell-free translation systems

FEBS Letters ◽  
1999 ◽  
Vol 462 (1-2) ◽  
pp. 43-46 ◽  
Author(s):  
Naoto Nemoto ◽  
Etsuko Miyamoto-Sato ◽  
Hiroshi Yanagawa
Keyword(s):  
Fluorescence Labeling ◽  
C Terminus ◽  
Free Translation ◽  
Translation Systems

scholarly journals Automated production of functional membrane proteins using eukaryotic cell-free translation systems

2015 ◽  
Vol 203 ◽  
pp. 45-53 ◽  
Author(s):  
Robert B. Quast ◽  
Oliver Kortt ◽  
Jörg Henkel ◽  
Srujan K. Dondapati ◽  
Doreen A. Wüstenhagen ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Eukaryotic Cell ◽  
Automated Production ◽  
Free Translation ◽  
Functional Membrane ◽  
Translation Systems
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