Q54. Which of the following enzyme(s), when overexpressed, would result in rice
grains with increased β–carotene content?
(A) Phytoene synthase
(B) Carotene desaturase
(C) β–glucoronidase
(D) Enolpyruvalshikimate–3–phosphate synthase (EPSPS)

Phytoene synthase and carotene desaturase overexpression lead to increased β-carotene in rice grains, as demonstrated in Golden Rice engineering.
Both options (A) and (B) are correct.

Option Analysis

Phytoene synthase (A): This enzyme catalyzes the first committed step in carotenoid biosynthesis, converting geranylgeranyl diphosphate (GGPP) to phytoene, the precursor to β-carotene. Overexpression, as in Golden Rice using daffodil or maize psy genes under endosperm-specific promoters, overcomes the rate-limiting step in rice endosperm, boosting flux toward β-carotene accumulation up to 37 μg/g in improved lines. Without it, no colored carotenoids form despite downstream enzymes.​

Carotene desaturase (B): This introduces double bonds into phytoene to form lycopene, which rice’s endogenous cyclases convert to β-carotene. Bacterial crtI (phytoene desaturase) from Erwinia/Pantoea, overexpressed in Golden Rice, bypasses missing plant desaturases (PDS, ZDS) and isomerases in endosperm, enabling lycopene production and subsequent β-carotene. Its activity ensures complete desaturation without phytoene buildup.​

β-glucuronidase (C): A reporter enzyme (gusA) used in plant transformation to visualize gene expression via histochemical assays; it hydrolyzes β-glucuronides but plays no role in carotenoid or β-carotene pathways. Overexpression yields blue color in assays, not golden grains.​

EPSPS (D): Enolpyruvylshikimate-3-phosphate synthase functions in the shikimate pathway for aromatic amino acids (phenylalanine, tyrosine, tryptophan), targeted by glyphosate herbicide. It provides no precursors or enzymes for carotenoid synthesis.​

Introduction
Golden Rice revolutionized biofortification by engineering rice grains with increased β-carotene content through targeted enzyme overexpression. This provitamin A enhancement combats vitamin A deficiency, vital for exams like CSIR NET Life Sciences. Key enzymes phytoene synthase and carotene desaturase drive the carotenoid pathway in rice endosperm.​

β-Carotene Pathway Basics

β-Carotene biosynthesis starts with GGPP condensation to phytoene by phytoene synthase (PSY), followed by desaturation to lycopene via carotene desaturases. Rice endosperm lacks PSY activity and efficient desaturases, halting the pathway despite downstream cyclases. Overexpression fills this gap, yielding yellow grains rich in β-carotene.​

Golden Rice Engineering

Two transgenes—psy (plant source) and crtI (bacterial carotene desaturase)—under endosperm promoters produce up to 31 μg/g β-carotene in GR2. PSY limits flux; maize psy boosts output 23-fold over originals. CRTI simplifies desaturation, avoiding multiple plant enzymes.​

Incorrect Options Explained

• β-glucuronidase: Reporter for transformation success, irrelevant to β-carotene.​

• EPSPS: Herbicide target in shikimate pathway, no carotenoid link.​

This biotech breakthrough highlights metabolic engineering for nutrition.​