NEIL1 Human

Nei Endonuclease VIII-Like 1 (NEIL1) is a human recombinant protein that plays a crucial role in the base excision repair (BER) pathway, which is responsible for repairing oxidative DNA damage. NEIL1 is a DNA glycosylase that recognizes and excises oxidized pyrimidines, such as thymine glycol, from DNA. This enzyme is essential for maintaining genomic stability and preventing mutations that can lead to various diseases, including cancer.

NEIL1 is a single, non-glycosylated polypeptide chain consisting of 410 amino acids and has a molecular mass of approximately 45.8 kDa . The protein is produced in Escherichia coli and is fused to a 20 amino acid His-tag at the N-terminus, which facilitates its purification using chromatographic techniques .

The crystal structure of NEIL1 reveals a unique “zincless finger” motif, which is required for its glycosylase activity . Unlike other members of the Fpg/Nei family that contain a zinc finger motif, NEIL1 has a structural motif composed of two antiparallel β-strands that mimic the zinc finger but do not coordinate zinc . This structural feature is essential for the enzyme’s ability to recognize and excise oxidized DNA lesions.

The recombinant NEIL1 protein is produced using an expression system in Escherichia coli. The gene encoding NEIL1 is cloned into an expression vector, which is then introduced into E. coli cells. The bacteria are cultured, and the expression of NEIL1 is induced. The protein is then purified using affinity chromatography, taking advantage of the His-tag at the N-terminus .

NEIL1 catalyzes the first step of the base excision repair pathway by recognizing and excising oxidized pyrimidines from DNA. The enzyme cleaves the N-glycosidic bond between the damaged base and the sugar-phosphate backbone, creating an apurinic/apyrimidinic (AP) site. This AP site is then processed by other enzymes in the BER pathway to complete the repair process .

The activity of NEIL1 can be analyzed using various biochemical assays. One common method is to incubate the enzyme with a DNA substrate containing an oxidized pyrimidine and then measure the release of the damaged base. The formation of AP sites can be detected using specific chemical reagents or by employing electrophoretic techniques to separate the cleaved DNA fragments .

NEIL1 is an important tool for studying the mechanisms of DNA repair and the role of oxidative damage in disease. Understanding how NEIL1 functions can provide insights into the development of therapeutic strategies for conditions associated with oxidative stress and genomic instability. Additionally, recombinant NEIL1 can be used in various research applications, including the development of assays for detecting DNA damage and the screening of potential DNA repair inhibitors.