# REGULATION OF PURINE BIOSYNTHESIS IN E COLI K12

> **NIH NIH R01** · SEATTLE BIOMEDICAL RESEARCH INSTITUTE · 1988 · $146,396

## Abstract

Seven genetic diseases have been identified to date that involve
alterations in purine metabolism.  These include Lesch-Nyhan
disease, severe combined immunodeficiency and most recently
discovered, a defect in adenylosuccinate lyase that results in
autism.  Understanding the regulation of purine metabolism will
aid in comprehending the basis of these diseases.  The object of
this proposed project is to understand the regulation of the de
novo purine biosynthetic pathway of Escherichia coli K12 on a
molecular basis.  The regulation of de novo purine biosynthesis has
three components:  (1) the promoter-control region where the
regulatory protein(s) bind; (2) the trans-acting regulatory
protein(s); and (3) the effector molecule(s).  The DNA sequence of
the purJHD and purB loci will be determined to complete the
sequence of the all known pur loci.  The control regions and
associated promoters will be identified by primer extension
studies and examined for the presence of the homologies and two
regions of imperfect dyad symmetry we have identified in the
other loci.  We will determine the role of these regions of dyad
symmetry in pur loci regulation by in vitro saturation mutagenesis
of selected control regions, i.e., purF, purA and guaBA loci.
After characterization of the control region mutations by Sanger
dideoxy sequencing, in vitro created pur-lac fusions will be
employed to assess the effect on the expression of individual
genes.  Mutations that result in decreased expression will be used
in a selection scheme to specifically identify trans-acting
regulatory loci.  This approach will distinguish between authentic
trans-acting regulatory mutations and effector molecule level
mutations which are normally genetically indistinguishable from
true regulatory mutations.  After isolation and identification, the
specific trans-acting regulatory mutations will be mapped with
linked transposons and standard recombinant DNA techniques will
be used to clone and sequence the corresponding locus.  The
regulation of the different loci of the pathway will be tested
under defined conditions in standard genetic background to
determined the relationship between the different control region
homologies and regulatory responses.

## Key facts

- **NIH application ID:** 3129585
- **Project number:** 2R01AI020068-06
- **Recipient organization:** SEATTLE BIOMEDICAL RESEARCH INSTITUTE
- **Principal Investigator:** JOHN M SMITH
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 1988
- **Award amount:** $146,396
- **Award type:** 2
- **Project period:** 1987-12-01 → 1991-11-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/3129585

## Citation

> US National Institutes of Health, RePORTER application 3129585, REGULATION OF PURINE BIOSYNTHESIS IN E COLI K12 (2R01AI020068-06). Retrieved via MORVS 2026-10-25 from https://api.morvs.ai/grant/nih/3129585. Licensed CC0.

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