Inhibition of serine palmitoyltransferase in vitro and long-chain base biosynthesis in intact Chinese hamster ovary cells by. beta. -chloroalanine

Journal Article · Tue Sep 06 04:00:00 EDT 1988 · Biochemistry; (United States)

DOI:https://doi.org/10.1021/bi00418a061· OSTI ID:6326428

Medlock, K A; Merrill, Jr, A H

The effects of ..beta..-chloroalanine (..beta..-Cl-alanine) on the serine palmitoyltransferase activity and the de novo biosynthesis of sphinganine and sphingenine were investigated in vitro with rat liver microsomes and in vivo with intact Chinese hamster ovary (CHO) cells. The inhibition in vitro was rapid, irreversible, and concentration and time dependent and apparently involved the active site because inactivation only occurred with ..beta..-Cl-L-alanine and was blocked by L-serine. These are characteristics of mechanism-based (suicide) inhibition. Serine palmitoyltransferase (SPT) was also inhibited when intact CHO cells were incubated with ..beta..-Cl-alanine and this treatment inhibited (/sup 14/C)serine incorporation into long-chain bases by intact cells. The concentration dependence of the loss of SPT activity and of long-chain base synthesis was identical. The effects of ..beta..-Cl-alanine appeared to occur with little perturbation of other cell functions: the cells exhibited no loss in cell viability, (/sup 14/C)serine uptake was not blocked, total lipid biosynthesis from (/sup 14/C)acetic acid was not decreased (nor was the appearance of radiolabel in cholesterol and phosphatidylcholine), and (/sup 3/H)thymidine incorporation into DNA was not affected. There appeared to be little effect on protein synthesis based on the incorporation of (/sup 3/H)leucine, which was only decreased by 14%. Although ..beta..-Cl-L-alanine is known to inhibit other pyridoxal 5'-phosphate dependent enzymes, alanine and aspartate transaminases were not inhibited under these conditions. These results establish the close association between the activity of serine palmitoyltransferase and the cellular rate of long-chain base formation and indicate that ..beta..-Cl-alanine and other mechanism-based inhibitors might be useful to study alterations in cellular long-chain base synthesis.

• Research Organization: Emory Univ., Atlanta, GA (USA)
• OSTI ID: 6326428
• Journal Information: Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 27:18; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ACETIC ACID
ALANINE-ALPHA
ALANINE-BETA
ALANINE-L
ALANINES
AMINO ACIDS
ANIMAL CELLS
ANIMALS
AZINES
BIOSYNTHESIS
BODY
CARBON 14 COMPOUNDS
CARBOXYLIC ACIDS
CHO CELLS
DIGESTIVE SYSTEM
ENZYMES
ESTERS
GLANDS
HETEROCYCLIC COMPOUNDS
HYDROXY ACIDS
IN VITRO
IN VIVO
INHIBITION
LABELLED COMPOUNDS
LEUCINE
LIPIDS
LIVER
MAMMALS
MONOCARBOXYLIC ACIDS
NUCLEOSIDES
NUCLEOTIDES
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
ORGANS
PHOSPHOLIPIDS
PYRIMIDINES
RATS
RIBOSIDES
RODENTS
SERINE
SYNTHESIS
THYMIDINE
TRANSFERASES
TRITIUM COMPOUNDS
UPTAKE
VERTEBRATES