Cloning and characterization of a second complementary DNA for human tryptase

Area of Science:

• Biochemistry
• Molecular Biology
• Genetics

Background:

• Human tryptase is a serine protease found in mast cells.
• Previous research identified alpha-tryptase cDNA.
• Understanding tryptase diversity is crucial for mast cell biology.

Purpose of the Study:

• To clone and characterize a second human tryptase cDNA, designated beta-tryptase.
• To compare beta-tryptase with the previously identified alpha-tryptase.
• To determine the genetic location and presence of both tryptase genes.

Main Methods:

• Cloning of beta-tryptase cDNA from a mast cell library.
• Nucleotide and amino acid sequencing of beta-tryptase.
• Comparison of alpha- and beta-tryptase sequences.
• Polymerase chain reaction (PCR) amplification of genomic DNA.
• Analysis of human/hamster somatic hybrids for gene localization.

Main Results:

• Beta-tryptase cDNA was successfully cloned and sequenced.
• Beta-tryptase shares 90% amino acid identity with alpha-tryptase, with 100% identity in the first 20 catalytic amino acids.
• Both alpha- and beta-tryptase sequences are present in the human genome and localized to chromosome 16.
• Minor differences in carbohydrate binding sites and substrate binding pockets were observed between alpha- and beta-tryptase.

Conclusions:

• The study validates the identity of both alpha- and beta-tryptase cDNA molecules.
• Both alpha- and beta-tryptase genes reside on human chromosome 16.
• This work provides a foundation for studying the differential expression and function of human tryptases.