Two regulatory models involving variable growth periods of β-galactosidase

Abstract

We have calculated the number of RNA polymerase molecules transcribing the induced lac operon of Escherichia coli as well as the distance between ribosomes on the proximal z and on the distal a messages†. These values were derived from: (a) rates of induced enzyme synthesis corrected by known turnover numbers to give numbers of enzyme monomers produced per cell per second; (b) the mass of each message derived either directly by hybridization of long-labeled RNA to specific DNA or from the rates of synthesis and decay obtained by hybridization of pulse-labeled RNA; the latter gives about 13 times better resolution; (c) conversion of message mass into a number of funtioning 3′ ends (producing finished polypeptides); size analyses indicate that the z message probably decays by a net directional degradation in the 5′ to 3′ direction. From this the fraction of completed molecules that are full-length and the total number with functional 3′ ends can be derived. (d) Rates of ribosome movement.Calculated values follow. (1) With a 3.3-second interval between transcription initiations, there are 38 molecules of RNA polymerase on z DNA per cell; with 1.7 copies of lac DNA, this gives 23 on each z cistron and five or six on the y and on the a cistrons. This corresponds to a spacing of about 135 nucleotides between polymerases, which is similar to that on the ribosomal cistrons which have only 1.7 seconds separating initiations. (2) There are 20 molecules of β-galactosidase monomer produced per cell per second from 1.1 × 10−4 pg of z RNA. 38% of the message molecules are nascent and not producing enzyme. Half of the completed molecules and 70% of their mass are intact z messages to give 62 molecules of z message containing functional 3′ ends/cell. Ribosomes load onto these messages at 3.2-second intervals to give a ribosomal spacing of 110 nucleotides. An average of 40 ribosomes translate each z message with half of the message molecules translated by more than 28 ribosomes and half by less. (3) Only 2.4 molecules of galactoside acetyltransferase monomer are produced per cell per second from 1.8 × 10−5 pg of a RNA. Only 20% of these molecules are nascent and 80% of the completed ones are intact; this gives 40 molecules of functioning a messages per cell. Ribosomes load to a message at 16-second intervals to give a spacing of 580 nucleotides.These results show that the frequencies of translation initiations can differ for different messages. The faster decay of a compared to z message is consistent with a model in which a ribosome can protect a vulnerable site near the start of a message from inactivation; messages that load less frequently decay faster. The combined net effect of these causally related processes could account for natural polarity (Zabin & Fowler, 1970), i.e. there is a fourfold lower production of enzyme from the a than from the z gene.